47 results on '"Beres, Stephen B."'
Search Results
2. Integrative Reverse Genetic Analysis Identifies Polymorphisms Contributing to Decreased Antimicrobial Agent Susceptibility in Streptococcus pyogenes .
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Beres SB, Zhu L, Pruitt L, Olsen RJ, Faili A, Kayal S, and Musser JM
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- Humans, Streptococcus pneumoniae genetics, Reverse Genetics, Penicillin-Binding Proteins genetics, beta-Lactams, Polymorphism, Genetic, Anti-Bacterial Agents pharmacology, Microbial Sensitivity Tests, beta-Lactam Resistance genetics, Streptococcus pyogenes genetics, Anti-Infective Agents
- Abstract
Identification of genetic polymorphisms causing increased antibiotic resistance in bacterial pathogens traditionally has proceeded from observed phenotype to defined mutant genotype. The availability of large collections of microbial genome sequences that lack antibiotic susceptibility metadata provides an important resource and opportunity to obtain new information about increased antimicrobial resistance by a reverse genotype-to-phenotype bioinformatic and experimental workflow. We analyzed 26,465 genome sequences of Streptococcus pyogenes, a human pathogen causing 700 million infections annually. The population genomic data identified amino acid changes in penicillin-binding proteins 1A, 1B, 2A, and 2X with signatures of evolution under positive selection as potential candidates for causing decreased susceptibility to β-lactam antibiotics. Construction and analysis of isogenic mutant strains containing individual amino acid replacements in penicillin-binding protein 2X (PBP2X) confirmed that the identified residues produced decreased susceptibility to penicillin. We also discovered the first chimeric PBP2X in S. pyogenes and show that strains containing it have significantly decreased β-lactam susceptibility. The novel integrative reverse genotype-to-phenotype strategy presented is broadly applicable to other pathogens and likely will lead to new knowledge about antimicrobial agent resistance, a massive public health problem worldwide. IMPORTANCE The recent demonstration that naturally occurring amino acid substitutions in Streptococcus pyogenes PBP2X are sufficient to cause severalfold reduced susceptibility to multiple β-lactam antibiotics in vitro raises the concern that these therapeutic agents may become compromised. Substitutions in PBP2X are common first-step mutations that, with the incremental accumulation of additional adaptive mutations within the PBPs, can result in high-level resistance. Because β-lactam susceptibility testing is not routinely performed, the nature and extent of such substitutions within the PBPs of S. pyogenes are poorly characterized. To address this knowledge deficit, polymorphisms in the PBPs were identified among the most comprehensive cohort of S. pyogenes genome sequences investigated to date. The mutational processes and selective forces acting on the PBPs were assessed to identify specific substitutions likely to influence β-lactam susceptibility and to evaluate factors posited to be impediments to resistance emergence.
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- 2022
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3. Population Genomic Molecular Epidemiological Study of Macrolide-Resistant Streptococcus pyogenes in Iceland, 1995 to 2016: Identification of a Large Clonal Population with a pbp2x Mutation Conferring Reduced In Vitro β-Lactam Susceptibility.
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Southon SB, Beres SB, Kachroo P, Saavedra MO, Erlendsdóttir H, Haraldsson G, Yerramilli P, Pruitt L, Zhu L, Musser JM, and Kristinsson KG
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- Anti-Bacterial Agents pharmacology, Drug Resistance, Bacterial, Epidemiologic Studies, Genotype, Humans, Iceland epidemiology, Macrolides pharmacology, Metagenomics, Microbial Sensitivity Tests, Mutation, beta-Lactams, Streptococcal Infections epidemiology, Streptococcus pyogenes genetics
- Abstract
Resistance to macrolide antibiotics is a global concern in the treatment of Streptococcus pyogenes (group A Streptococcus [GAS]) infections. In Iceland, since the detection of the first macrolide-resistant isolate in 1998, three epidemic waves of macrolide-resistant GAS infections have occurred, with peaks in 1999, 2004, and 2008. We conducted whole-genome sequencing of all 1,575 available GAS macrolide-resistant clinical isolates of all infection types collected at the national reference laboratory in Reykjavik, Iceland, from 1998 to 2016. Among 1,515 erythromycin-resistant isolates, 90.3% were of only three emm types, emm4 ( n = 713), emm6 ( n = 324), and emm12 ( n = 332), with each being predominant in a distinct epidemic peak. The antibiotic efflux pump genes, mef (A) and msr (D), were present on chimeric mobile genetic elements in 99.3% of the macrolide-resistant isolates of these emm types. Of note, in addition to macrolide resistance, virtually all emm12 isolates had a single amino acid substitution in penicillin-binding protein PBP2X that conferred a 2-fold increased penicillin G and ampicillin MIC among the isolates tested. We conclude that each of the three large epidemic peaks of macrolide-resistant GAS infections occurring in Iceland since 1998 was caused by the emergence and clonal expansion of progenitor strains, with macrolide resistance being conferred predominantly by inducible Mef(A) and Msr(D) drug efflux pumps. The occurrence of emm12 strains with macrolide resistance and decreased beta-lactam susceptibility was unexpected and is of public health concern., (Copyright © 2020 Southon et al.)
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- 2020
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4. Streptococcus pyogenes genes that promote pharyngitis in primates.
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Zhu L, Olsen RJ, Beres SB, Saavedra MO, Kubiak SL, Cantu CC, Jenkins L, Waller AS, Sun Z, Palzkill T, Porter AR, DeLeo FR, and Musser JM
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- Animals, Disease Models, Animal, Genome-Wide Association Study, Humans, Macaca fascicularis, Genes, Bacterial, Pharyngitis genetics, Pharyngitis metabolism, Pharyngitis microbiology, Pharyngitis pathology, Streptococcal Infections genetics, Streptococcal Infections metabolism, Streptococcal Infections pathology, Streptococcus pyogenes genetics, Streptococcus pyogenes metabolism, Streptococcus pyogenes pathogenicity, Virulence Factors genetics, Virulence Factors metabolism
- Abstract
Streptococcus pyogenes (group A streptococcus; GAS) causes 600 million cases of pharyngitis annually worldwide. There is no licensed human GAS vaccine despite a century of research. Although the human oropharynx is the primary site of GAS infection, the pathogenic genes and molecular processes used to colonize, cause disease, and persist in the upper respiratory tract are poorly understood. Using dense transposon mutant libraries made with serotype M1 and M28 GAS strains and transposon-directed insertion sequencing, we performed genome-wide screens in the nonhuman primate (NHP) oropharynx. We identified many potentially novel GAS fitness genes, including a common set of 115 genes that contribute to fitness in both genetically distinct GAS strains during experimental NHP pharyngitis. Targeted deletion of 4 identified fitness genes/operons confirmed that our newly identified targets are critical for GAS virulence during experimental pharyngitis. Our screens discovered many surface-exposed or secreted proteins - substrates for vaccine research - that potentially contribute to GAS pharyngitis, including lipoprotein HitA. Pooled human immune globulin reacted with purified HitA, suggesting that humans produce antibodies against this lipoprotein. Our findings provide new information about GAS fitness in the upper respiratory tract that may assist in translational research, including developing novel vaccines.
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- 2020
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5. Genome-Wide Screens Identify Group A Streptococcus Surface Proteins Promoting Female Genital Tract Colonization and Virulence.
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Zhu L, Olsen RJ, Beres SB, Ojeda Saavedra M, Kubiak SL, Cantu CC, Jenkins L, Yerramilli P, Pruitt L, Charbonneau ARL, Waller AS, and Musser JM
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- Animals, Bacterial Proteins genetics, Disease Models, Animal, Female, Gene Expression Regulation, Bacterial, Macaca fascicularis, Membrane Proteins genetics, Streptococcal Infections metabolism, Streptococcus pyogenes metabolism, Vaginal Diseases pathology, Virulence, Bacterial Proteins metabolism, Membrane Proteins metabolism, Streptococcal Infections microbiology, Streptococcus pyogenes genetics, Streptococcus pyogenes pathogenicity, Vaginal Diseases microbiology
- Abstract
Group A streptococcus (GAS) is a major pathogen that impacts health and economic affairs worldwide. Although the oropharynx is the primary site of infection, GAS can colonize the female genital tract and cause severe diseases, such as puerperal sepsis, neonatal infections, and necrotizing myometritis. Our understanding of how GAS genes contribute to interaction with the primate female genital tract is limited by the lack of relevant animal models. Using two genome-wide transposon mutagenesis screens, we identified 69 GAS genes required for colonization of the primate vaginal mucosa in vivo and 96 genes required for infection of the uterine wall ex vivo. We discovered a common set of 39 genes important for GAS fitness in both environments. They include genes encoding transporters, surface proteins, transcriptional regulators, and metabolic pathways. Notably, the genes that encode the surface-exclusion protein (SpyAD) and the immunogenic secreted protein 2 (Isp2) were found to be crucial for GAS fitness in the female primate genital tract. Targeted gene deletion confirmed that isogenic mutant strains ΔspyAD and Δisp2 are significantly impaired in ability to colonize the primate genital tract and cause uterine wall pathologic findings. Our studies identified novel GAS genes that contribute to female reproductive tract interaction that warrant translational research investigation., (Copyright © 2020 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)
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- 2020
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6. Genetic heterogeneity of the Spy1336/R28-Spy1337 virulence axis in Streptococcus pyogenes and effect on gene transcript levels and pathogenesis.
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Eraso JM, Kachroo P, Olsen RJ, Beres SB, Zhu L, Badu T, Shannon S, Cantu CC, Saavedra MO, Kubiak SL, Porter AR, DeLeo FR, and Musser JM
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- Animals, Disease Models, Animal, Fasciitis, Necrotizing pathology, Gene Expression Regulation, Bacterial, Genetic Heterogeneity, Humans, Mice, Polymorphism, Genetic, Streptococcal Infections pathology, Transcriptome, Virulence Factors genetics, Bacterial Proteins genetics, Fasciitis, Necrotizing microbiology, Streptococcal Infections microbiology, Streptococcus pyogenes genetics, Streptococcus pyogenes isolation & purification, Virulence genetics
- Abstract
Streptococcus pyogenes is a strict human pathogen responsible for more than 700 million infections annually worldwide. Strains of serotype M28 S. pyogenes are typically among the five more abundant types causing invasive infections and pharyngitis in adults and children. Type M28 strains also have an unusual propensity to cause puerperal sepsis and neonatal disease. We recently discovered that a one-nucleotide indel in an intergenic homopolymeric tract located between genes Spy1336/R28 and Spy1337 altered virulence in a mouse model of infection. In the present study, we analyzed size variation in this homopolymeric tract and determined the extent of heterogeneity in the number of tandemly-repeated 79-amino acid domains in the coding region of Spy1336/R28 in large samples of strains recovered from humans with invasive infections. Both repeat sequence elements are highly polymorphic in natural populations of M28 strains. Variation in the homopolymeric tract results in (i) changes in transcript levels of Spy1336/R28 and Spy1337 in vitro, (ii) differences in virulence in a mouse model of necrotizing myositis, and (iii) global transcriptome changes as shown by RNAseq analysis of isogenic mutant strains. Variation in the number of tandem repeats in the coding sequence of Spy1336/R28 is responsible for size variation of R28 protein in natural populations. Isogenic mutant strains in which genes encoding R28 or transcriptional regulator Spy1337 are inactivated are significantly less virulent in a nonhuman primate model of necrotizing myositis. Our findings provide impetus for additional studies addressing the role of R28 and Spy1337 variation in pathogen-host interactions., Competing Interests: The authors have declared that no competing interests exist.
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- 2020
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7. Reduced In Vitro Susceptibility of Streptococcus pyogenes to β-Lactam Antibiotics Associated with Mutations in the pbp2x Gene Is Geographically Widespread.
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Musser JM, Beres SB, Zhu L, Olsen RJ, Vuopio J, Hyyryläinen HL, Gröndahl-Yli-Hannuksela K, Kristinsson KG, Darenberg J, Henriques-Normark B, Hoffmann S, Caugant DA, Smith AJ, Lindsay DSJ, Boragine DM, and Palzkill T
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- Anti-Bacterial Agents pharmacology, Bacterial Proteins genetics, Humans, Microbial Sensitivity Tests, Mutation, Penicillin-Binding Proteins genetics, Phylogeny, Streptococcus pyogenes genetics, beta-Lactams pharmacology
- Abstract
Recently, two related Streptococcus pyogenes strains with reduced susceptibility to ampicillin, amoxicillin, and cefotaxime, antibiotics commonly used to treat S. pyogenes infections, were reported. The two strains had the same nonsynonymous (amino acid-substituting) mutation in the pbp2x gene, encoding penicillin-binding protein 2X (PBP2X). This concerning report led us to investigate our library of 7,025 genome sequences of type emm1 , emm28 , and emm89 S. pyogenes clinical strains recovered from intercontinental sources for mutations in pbp2x We identified 137 strains that, combined, had 37 nonsynonymous mutations in 36 codons in pbp2x Although to a lesser magnitude than the two previously published isolates, many of our strains had decreased susceptibility in vitro to multiple beta-lactam antibiotics. Many pbp2x mutations were found only in single strains, but 16 groups of two or more isolates of the same emm type had an identical amino acid replacement. Phylogenetic analysis showed that, with one exception, strains of the same emm type with the same amino acid replacement were clonally related by descent. This finding indicates that strains with some amino acid changes in PBP2X can successfully spread to new human hosts and cause invasive infections. Mapping of the amino acid changes onto a three-dimensional structure of the related Streptococcus pneumoniae PBP2X suggests that some substitutions are located in regions functionally important in related pathogenic bacterial species. Decreased beta-lactam susceptibility is geographically widespread in strains of numerically common emm gene subtypes. Enhanced surveillance and further epidemiological and molecular genetic study of this potential emergent antimicrobial problem are warranted., (Copyright © 2020 American Society for Microbiology.)
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- 2020
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8. Integrated analysis of population genomics, transcriptomics and virulence provides novel insights into Streptococcus pyogenes pathogenesis.
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Kachroo P, Eraso JM, Beres SB, Olsen RJ, Zhu L, Nasser W, Bernard PE, Cantu CC, Saavedra MO, Arredondo MJ, Strope B, Do H, Kumaraswami M, Vuopio J, Gröndahl-Yli-Hannuksela K, Kristinsson KG, Gottfredsson M, Pesonen M, Pensar J, Davenport ER, Clark AG, Corander J, Caugant DA, Gaini S, Magnussen MD, Kubiak SL, Nguyen HAT, Long SW, Porter AR, DeLeo FR, and Musser JM
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- Gene Expression Regulation, Bacterial genetics, Genome-Wide Association Study methods, Genomics methods, Phylogeny, Quantitative Trait Loci genetics, Genome, Bacterial genetics, Streptococcus pyogenes genetics, Transcriptome genetics, Virulence genetics
- Abstract
Streptococcus pyogenes causes 700 million human infections annually worldwide, yet, despite a century of intensive effort, there is no licensed vaccine against this bacterium. Although a number of large-scale genomic studies of bacterial pathogens have been published, the relationships among the genome, transcriptome, and virulence in large bacterial populations remain poorly understood. We sequenced the genomes of 2,101 emm28 S. pyogenes invasive strains, from which we selected 492 phylogenetically diverse strains for transcriptome analysis and 50 strains for virulence assessment. Data integration provided a novel understanding of the virulence mechanisms of this model organism. Genome-wide association study, expression quantitative trait loci analysis, machine learning, and isogenic mutant strains identified and confirmed a one-nucleotide indel in an intergenic region that significantly alters global transcript profiles and ultimately virulence. The integrative strategy that we used is generally applicable to any microbe and may lead to new therapeutics for many human pathogens.
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- 2019
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9. Gene fitness landscape of group A streptococcus during necrotizing myositis.
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Zhu L, Olsen RJ, Beres SB, Eraso JM, Saavedra MO, Kubiak SL, Cantu CC, Jenkins L, Charbonneau ARL, Waller AS, and Musser JM
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- Animals, Cell Line, Disease Models, Animal, Humans, Mice, Bacterial Proteins biosynthesis, Bacterial Proteins genetics, Carrier Proteins biosynthesis, Carrier Proteins genetics, Fasciitis, Necrotizing genetics, Fasciitis, Necrotizing metabolism, Fasciitis, Necrotizing pathology, Gene Expression Regulation, Bacterial, Myositis genetics, Myositis metabolism, Myositis microbiology, Myositis pathology, Streptococcus pyogenes genetics, Streptococcus pyogenes metabolism, Streptococcus pyogenes pathogenicity
- Abstract
Necrotizing fasciitis and myositis are devastating infections characterized by high mortality. Group A streptococcus (GAS) is a common cause of these infections, but the molecular pathogenesis is poorly understood. We report a genome-wide analysis using serotype M1 and M28 strains that identified GAS genes contributing to necrotizing myositis in nonhuman primates (NHP), a clinically relevant model. Using transposon-directed insertion-site sequencing (TraDIS), we identified 126 and 116 GAS genes required for infection by serotype M1 and M28 organisms, respectively. For both M1 and M28 strains, more than 25% of the GAS genes required for necrotizing myositis encode known or putative transporters. Thirteen GAS transporters contributed to both M1 and M28 strain fitness in NHP myositis, including putative importers for amino acids, carbohydrates, and vitamins and exporters for toxins, quorum-sensing peptides, and uncharacterized molecules. Targeted deletion of genes encoding 5 transporters confirmed that each isogenic mutant strain was significantly (P < 0.05) impaired in causing necrotizing myositis in NHPs. Quantitative reverse-transcriptase PCR (qRT-PCR) analysis showed that these 5 genes are expressed in infected NHP and human skeletal muscle. Certain substrate-binding lipoproteins of these transporters, such as Spy0271 and Spy1728, were previously documented to be surface exposed, suggesting that our findings have translational research implications.
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- 2019
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10. RocA Has Serotype-Specific Gene Regulatory and Pathogenesis Activities in Serotype M28 Group A Streptococcus.
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Bernard PE, Kachroo P, Zhu L, Beres SB, Eraso JM, Kajani Z, Long SW, Musser JM, and Olsen RJ
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- Animals, Colony-Forming Units Assay, Disease Models, Animal, Gene Deletion, Gene Expression Profiling, Mice, Myositis microbiology, Streptococcal Infections microbiology, Streptococcus pyogenes growth & development, Trans-Activators genetics, Virulence Factors biosynthesis, Virulence Factors genetics, Gene Expression Regulation, Bacterial, Myositis pathology, Streptococcal Infections pathology, Streptococcus pyogenes genetics, Streptococcus pyogenes pathogenicity, Trans-Activators metabolism
- Abstract
Serotype M28 group A streptococcus (GAS) is a common cause of infections such as pharyngitis ("strep throat") and necrotizing fasciitis ("flesh-eating" disease). Relatively little is known about the molecular mechanisms underpinning M28 GAS pathogenesis. Whole-genome sequencing studies of M28 GAS strains recovered from patients with invasive infections found an unexpectedly high number of missense (amino acid-changing) and nonsense (protein-truncating) polymorphisms in rocA ( r egulator o f C ov), leading us to hypothesize that altered RocA activity contributes to M28 GAS molecular pathogenesis. To test this hypothesis, an isogenic rocA deletion mutant strain was created. Transcriptome sequencing (RNA-seq) analysis revealed that RocA inactivation significantly alters the level of transcripts for 427 and 323 genes at mid-exponential and early stationary growth phases, respectively, including genes for 41 transcription regulators and 21 virulence factors. In contrast, RocA transcriptomes from other GAS M protein serotypes are much smaller and include fewer transcription regulators. The rocA mutant strain had significantly increased secreted activity of multiple virulence factors and grew to significantly higher colony counts under acid stress in vitro RocA inactivation also significantly increased GAS virulence in a mouse model of necrotizing myositis. Our results demonstrate that RocA is an important regulator of transcription regulators and virulence factors in M28 GAS and raise the possibility that naturally occurring polymorphisms in rocA in some fashion contribute to human invasive infections caused by M28 GAS strains., (Copyright © 2018 American Society for Microbiology.)
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- 2018
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11. Genome sequence analysis of emm89 Streptococcus pyogenes strains causing infections in Scotland, 2010-2016.
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Beres SB, Olsen RJ, Ojeda Saavedra M, Ure R, Reynolds A, Lindsay DSJ, Smith AJ, and Musser JM
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- DNA, Bacterial genetics, Humans, Molecular Epidemiology, Retrospective Studies, Scotland epidemiology, Sequence Analysis, DNA, Streptococcus pyogenes classification, Streptococcus pyogenes isolation & purification, Bacterial Proteins genetics, Genome, Bacterial, Streptococcal Infections epidemiology, Streptococcus pyogenes genetics
- Abstract
Purpose: Strains of type emm89 Streptococcus pyogenes have recently increased in frequency as a cause of human infections in several countries in Europe and North America. This increase has been molecular epidemiologically linked with the emergence of a new genetically distinct clone, designated clade 3. We sought to extend our understanding of this epidemic behavior by the genetic characterization of type emm89 strains responsible in recent years for an increased frequency of infections in Scotland., Methodology: We sequenced the genomes of a retrospective cohort of 122 emm89 strains recovered from patients with invasive and noninvasive infections throughout Scotland during 2010 to 2016., Results: All but one of the 122 emm89 infection isolates are of the recently emerged epidemic clade 3 clonal lineage. The Scotland isolates are closely related to and not genetically distinct from recent emm89 strains from England, they constitute a single genetic population., Conclusions: The clade 3 clone causes virtually all-contemporary emm89 infections in Scotland. These findings add Scotland to a growing list of countries of Europe and North America where, by whole genome sequencing, emm89 clade 3 strains have been demonstrated to be the cause of an ongoing epidemic of invasive infections and to be genetically related due to descent from a recent common progenitor.
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- 2017
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12. Interacting networks of resistance, virulence and core machinery genes identified by genome-wide epistasis analysis.
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Skwark MJ, Croucher NJ, Puranen S, Chewapreecha C, Pesonen M, Xu YY, Turner P, Harris SR, Beres SB, Musser JM, Parkhill J, Bentley SD, Aurell E, and Corander J
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- Aminoacyltransferases genetics, Anti-Bacterial Agents therapeutic use, Bacterial Proteins genetics, Gene Regulatory Networks genetics, Genetics, Population, Genome, Bacterial genetics, Genomics, Genotype, Humans, Microbial Sensitivity Tests, Penicillin-Binding Proteins chemistry, Penicillin-Binding Proteins genetics, Peptidyl Transferases genetics, Streptococcus pneumoniae drug effects, Streptococcus pneumoniae pathogenicity, Streptococcus pyogenes drug effects, Streptococcus pyogenes pathogenicity, beta-Lactams metabolism, Epistasis, Genetic, Selection, Genetic genetics, Streptococcus pneumoniae genetics, Streptococcus pyogenes genetics, beta-Lactam Resistance genetics
- Abstract
Recent advances in the scale and diversity of population genomic datasets for bacteria now provide the potential for genome-wide patterns of co-evolution to be studied at the resolution of individual bases. Here we describe a new statistical method, genomeDCA, which uses recent advances in computational structural biology to identify the polymorphic loci under the strongest co-evolutionary pressures. We apply genomeDCA to two large population data sets representing the major human pathogens Streptococcus pneumoniae (pneumococcus) and Streptococcus pyogenes (group A Streptococcus). For pneumococcus we identified 5,199 putative epistatic interactions between 1,936 sites. Over three-quarters of the links were between sites within the pbp2x, pbp1a and pbp2b genes, the sequences of which are critical in determining non-susceptibility to beta-lactam antibiotics. A network-based analysis found these genes were also coupled to that encoding dihydrofolate reductase, changes to which underlie trimethoprim resistance. Distinct from these antibiotic resistance genes, a large network component of 384 protein coding sequences encompassed many genes critical in basic cellular functions, while another distinct component included genes associated with virulence. The group A Streptococcus (GAS) data set population represents a clonal population with relatively little genetic variation and a high level of linkage disequilibrium across the genome. Despite this, we were able to pinpoint two RNA pseudouridine synthases, which were each strongly linked to a separate set of loci across the chromosome, representing biologically plausible targets of co-selection. The population genomic analysis method applied here identifies statistically significantly co-evolving locus pairs, potentially arising from fitness selection interdependence reflecting underlying protein-protein interactions, or genes whose product activities contribute to the same phenotype. This discovery approach greatly enhances the future potential of epistasis analysis for systems biology, and can complement genome-wide association studies as a means of formulating hypotheses for targeted experimental work.
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- 2017
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13. Genomic Characteristics Behind the Spread of Bacteremic Group A Streptococcus Type emm89 in Finland, 2004-2014.
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Latronico F, Nasser W, Puhakainen K, Ollgren J, Hyyryläinen HL, Beres SB, Lyytikäinen O, Jalava J, Musser JM, and Vuopio J
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- Adolescent, Adult, Aged, Aged, 80 and over, Antigens, Bacterial genetics, Bacteremia mortality, Bacterial Outer Membrane Proteins genetics, Carrier Proteins genetics, Child, Child, Preschool, Cluster Analysis, Female, Finland epidemiology, Genome, Bacterial, Genomics, Humans, Infant, Infant, Newborn, Male, Middle Aged, Molecular Diagnostic Techniques methods, Molecular Epidemiology methods, Phylogeny, Polymerase Chain Reaction methods, Streptococcal Infections mortality, Streptococcus pyogenes isolation & purification, Survival Analysis, Young Adult, Bacteremia epidemiology, Bacteremia microbiology, Genotype, Streptococcal Infections epidemiology, Streptococcal Infections microbiology, Streptococcus pyogenes classification, Streptococcus pyogenes genetics
- Abstract
Background: Many countries worldwide have reported increasing numbers of emm89 group A Streptococcus (GAS) infections during last decade. Pathogen genetic factors linked to this increase need assessment., Methods: We investigated epidemiological characteristics of emm89 GAS bacteremic infections, including 7-day and 30-day case-fatality rates, in Finland during 2004-2014 and linked them to whole-genome sequencing data obtained from corresponding strains. The Fisher exact test and exact logistic regression were used to compare differences between bacteremic infections due to emm89 GAS belonging to different genetic clades and subclades., Results: Out of 1928 cases of GAS bacteremic infection, 278 were caused by emm89 GAS. We identified 2 genetically distinct clades, arbitrarily designated clade 2 and clade 3. Both clades were present during 2004-2008, but clade 3 increased rapidly from 2009 onward. Six subclades (designated subclades A-F) were identified within clade 3, based on phylogenetic core genome analysis. The case-fatality rate differed significantly between subclades (P < .05), with subclade D having the highest 30-day estimated case-fatality rate (19% vs 3%-14%)., Conclusions: A new emm89 clone, clade 3, emerged in 2009 and spread rapidly in Finland. Patients infected with certain subclades of clade 3 were significantly more likely to die. A specific polymerase chain reaction assay was developed to follow the spread of subclade D in 2015., (© The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America.)
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- 2016
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14. Genomic Landscape of Intrahost Variation in Group A Streptococcus: Repeated and Abundant Mutational Inactivation of the fabT Gene Encoding a Regulator of Fatty Acid Synthesis.
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Eraso JM, Olsen RJ, Beres SB, Kachroo P, Porter AR, Nasser W, Bernard PE, DeLeo FR, and Musser JM
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- Amino Acid Sequence, Animals, Bacterial Proteins genetics, Base Sequence, DNA, Bacterial genetics, Fasciitis, Necrotizing microbiology, Gene Expression Regulation, Bacterial, Host Specificity, Macaca fascicularis, Mutation, Polymorphism, Genetic, Bacterial Proteins metabolism, Fatty Acids biosynthesis, Streptococcus pyogenes genetics
- Abstract
To obtain new information about Streptococcus pyogenes intrahost genetic variation during invasive infection, we sequenced the genomes of 2,954 serotype M1 strains recovered from a nonhuman primate experimental model of necrotizing fasciitis. A total of 644 strains (21.8%) acquired polymorphisms relative to the input parental strain. The fabT gene, encoding a transcriptional regulator of fatty acid biosynthesis genes, contained 54.5% of these changes. The great majority of polymorphisms were predicted to deleteriously alter FabT function. Transcriptome-sequencing (RNA-seq) analysis of a wild-type strain and an isogenic fabT deletion mutant strain found that between 3.7 and 28.5% of the S. pyogenes transcripts were differentially expressed, depending on the growth temperature (35°C or 40°C) and growth phase (mid-exponential or stationary phase). Genes implicated in fatty acid synthesis and lipid metabolism were significantly upregulated in the fabT deletion mutant strain. FabT also directly or indirectly regulated central carbon metabolism genes, including pyruvate hub enzymes and fermentation pathways and virulence genes. Deletion of fabT decreased virulence in a nonhuman primate model of necrotizing fasciitis. In addition, the fabT deletion strain had significantly decreased survival in human whole blood and during phagocytic interaction with polymorphonuclear leukocytes ex vivo We conclude that FabT mutant progeny arise during infection, constitute a metabolically distinct subpopulation, and are less virulent in the experimental models used here., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)
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- 2016
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15. Transcriptome Remodeling Contributes to Epidemic Disease Caused by the Human Pathogen Streptococcus pyogenes.
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Beres SB, Kachroo P, Nasser W, Olsen RJ, Zhu L, Flores AR, de la Riva I, Paez-Mayorga J, Jimenez FE, Cantu C, Vuopio J, Jalava J, Kristinsson KG, Gottfredsson M, Corander J, Fittipaldi N, Di Luca MC, Petrelli D, Vitali LA, Raiford A, Jenkins L, and Musser JM
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- Evolution, Molecular, Gene Transfer, Horizontal, Genome, Bacterial, Genotype, Humans, Immune Evasion, Polymorphism, Single Nucleotide, Recombination, Genetic, Streptococcus pyogenes immunology, Streptococcus pyogenes pathogenicity, Virulence genetics, Virulence Factors genetics, Bacterial Proteins genetics, Epidemics prevention & control, Host-Pathogen Interactions, Streptococcal Infections epidemiology, Streptococcal Infections microbiology, Streptococcus pyogenes genetics, Transcriptome
- Abstract
Unlabelled: For over a century, a fundamental objective in infection biology research has been to understand the molecular processes contributing to the origin and perpetuation of epidemics. Divergent hypotheses have emerged concerning the extent to which environmental events or pathogen evolution dominates in these processes. Remarkably few studies bear on this important issue. Based on population pathogenomic analysis of 1,200 Streptococcus pyogenes type emm89 infection isolates, we report that a series of horizontal gene transfer events produced a new pathogenic genotype with increased ability to cause infection, leading to an epidemic wave of disease on at least two continents. In the aggregate, these and other genetic changes substantially remodeled the transcriptomes of the evolved progeny, causing extensive differential expression of virulence genes and altered pathogen-host interaction, including enhanced immune evasion. Our findings delineate the precise molecular genetic changes that occurred and enhance our understanding of the evolutionary processes that contribute to the emergence and persistence of epidemically successful pathogen clones. The data have significant implications for understanding bacterial epidemics and for translational research efforts to blunt their detrimental effects., Importance: The confluence of studies of molecular events underlying pathogen strain emergence, evolutionary genetic processes mediating altered virulence, and epidemics is in its infancy. Although understanding these events is necessary to develop new or improved strategies to protect health, surprisingly few studies have addressed this issue, in particular, at the comprehensive population genomic level. Herein we establish that substantial remodeling of the transcriptome of the human-specific pathogen Streptococcus pyogenes by horizontal gene flow and other evolutionary genetic changes is a central factor in precipitating and perpetuating epidemic disease. The data unambiguously show that the key outcome of these molecular events is evolution of a new, more virulent pathogenic genotype. Our findings provide new understanding of epidemic disease., (Copyright © 2016 Beres et al.)
- Published
- 2016
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16. The majority of 9,729 group A streptococcus strains causing disease secrete SpeB cysteine protease: pathogenesis implications.
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Olsen RJ, Raghuram A, Cantu C, Hartman MH, Jimenez FE, Lee S, Ngo A, Rice KA, Saddington D, Spillman H, Valson C, Flores AR, Beres SB, Long SW, Nasser W, and Musser JM
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- Animals, Bacterial Proteins metabolism, Caseins chemistry, Epidemiological Monitoring, Europe epidemiology, Exotoxins metabolism, Fasciitis, Necrotizing epidemiology, Fasciitis, Necrotizing microbiology, Fasciitis, Necrotizing pathology, High-Throughput Nucleotide Sequencing, Humans, Mutation, Pharyngitis epidemiology, Pharyngitis microbiology, Pharyngitis pathology, Primates, Proteolysis, Serotyping, Streptococcal Infections epidemiology, Streptococcal Infections microbiology, Streptococcal Infections pathology, Streptococcus pyogenes classification, Streptococcus pyogenes pathogenicity, Transcription, Genetic, United States epidemiology, Virulence, Bacterial Proteins genetics, Exotoxins genetics, Gene Expression Regulation, Bacterial, Genome, Bacterial, Streptococcus pyogenes enzymology, Streptococcus pyogenes genetics
- Abstract
Group A streptococcus (GAS), the causative agent of pharyngitis and necrotizing fasciitis, secretes the potent cysteine protease SpeB. Several lines of evidence suggest that SpeB is an important virulence factor. SpeB is expressed in human infections, protects mice from lethal challenge when used as a vaccine, and contributes significantly to tissue destruction and dissemination in animal models. However, recent descriptions of mutations in genes implicated in SpeB production have led to the idea that GAS may be under selective pressure to decrease secreted SpeB protease activity during infection. Thus, two divergent hypotheses have been proposed. One postulates that SpeB is a key contributor to pathogenesis; the other, that GAS is under selection to decrease SpeB during infection. In order to distinguish between these alternative hypotheses, we performed casein hydrolysis assays to measure the SpeB protease activity secreted by 6,775 GAS strains recovered from infected humans. The results demonstrated that 84.3% of the strains have a wild-type SpeB protease phenotype. The availability of whole-genome sequence data allowed us to determine the relative frequencies of mutations in genes implicated in SpeB production. The most abundantly mutated genes were direct transcription regulators. We also sequenced the genomes of 2,954 GAS isolates recovered from nonhuman primates with experimental necrotizing fasciitis. No mutations that would result in a SpeB-deficient phenotype were identified. Taken together, these data unambiguously demonstrate that the great majority of GAS strains recovered from infected humans secrete wild-type levels of SpeB protease activity. Our data confirm the important role of SpeB in GAS pathogenesis and help end a long-standing controversy., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)
- Published
- 2015
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17. A molecular trigger for intercontinental epidemics of group A Streptococcus.
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Zhu L, Olsen RJ, Nasser W, Beres SB, Vuopio J, Kristinsson KG, Gottfredsson M, Porter AR, DeLeo FR, and Musser JM
- Subjects
- Bacterial Proteins, Fasciitis, Necrotizing epidemiology, Fasciitis, Necrotizing genetics, Humans, Leukocytes microbiology, Pharyngitis epidemiology, Pharyngitis genetics, Streptolysins, NAD+ Nucleosidase genetics, Pandemics, Polymorphism, Genetic, Streptococcal Infections epidemiology, Streptococcal Infections genetics, Streptococcus pyogenes genetics, Streptococcus pyogenes pathogenicity, Virulence Factors genetics
- Abstract
The identification of the molecular events responsible for strain emergence, enhanced virulence, and epidemicity has been a long-pursued goal in infectious diseases research. A recent analysis of 3,615 genomes of serotype M1 group A Streptococcus strains (the so-called "flesh-eating" bacterium) identified a recombination event that coincides with the global M1 pandemic beginning in the early 1980s. Here, we have shown that the allelic variation that results from this recombination event, which replaces the chromosomal region encoding secreted NADase and streptolysin O, is the key driver of increased toxin production and enhanced infection severity of the M1 pandemic strains. Using isoallelic mutant strains, we found that 3 polymorphisms in this toxin gene region increase resistance to killing by human polymorphonuclear leukocytes, increase bacterial proliferation, and increase virulence in animal models of pharyngitis and necrotizing fasciitis. Genome sequencing of an additional 1,125 streptococcal strains and virulence studies revealed that a highly similar recombinational replacement event underlies an ongoing intercontinental epidemic of serotype M89 group A Streptococcus infections. By identifying the molecular changes that enhance upper respiratory tract fitness, increased resistance to innate immunity, and increased tissue destruction, we describe a mechanism that underpins epidemic streptococcal infections, which have affected many millions of people.
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- 2015
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18. Clinical laboratory response to a mock outbreak of invasive bacterial infections: a preparedness study.
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Olsen RJ, Fittipaldi N, Kachroo P, Sanson MA, Long SW, Como-Sabetti KJ, Valson C, Cantu C, Lynfield R, Van Beneden C, Beres SB, and Musser JM
- Subjects
- Animals, Canada, Disease Models, Animal, Gene Expression Profiling, Genome, Bacterial, Health Services Research, High-Throughput Nucleotide Sequencing methods, Humans, Mice, Molecular Diagnostic Techniques methods, Streptococcus pyogenes classification, Streptococcus pyogenes genetics, United States, Civil Defense, Disease Outbreaks, Laboratory Proficiency Testing, Streptococcal Infections diagnosis, Streptococcal Infections epidemiology, Streptococcus pyogenes isolation & purification
- Abstract
Large hospital-based clinical laboratories must be prepared to rapidly investigate potential infectious disease outbreaks. To challenge the ability of our molecular diagnostics laboratory to use whole-genome sequencing in a potential outbreak scenario and identify impediments to these efforts, we studied 84 invasive serotype emm59 group A streptococcus (GAS) strains collected in the United States. We performed a rapid-response exercise to the mock outbreak scenario using whole-genome sequencing, genome-wide transcript analysis, and mouse virulence studies. The protocol changes installed in response to the lessons learned were tested in a second iteration. The initial investigation was completed in 9 days. Whole-genome sequencing showed that the invasive infections were caused by multiple subclones of epidemic emm59 GAS strains likely spread to the United States from Canada. The phylogenetic tree showed a strong temporal-spatial structure with diversity in mobile genetic element content, features that are useful for identifying closely related strains and possible transmission events. The genome data informed the epidemiology, identifying multiple patients who likely acquired the organisms through direct person-to-person transmission. Transcriptome analysis unexpectedly revealed significantly altered expression of genes encoding a two-component regulator and the hyaluronic acid capsule virulence factor. Mouse infection studies confirmed a high-virulence capacity of these emm59 organisms. Whole-genome sequencing, coupled with transcriptome analysis and animal virulence studies, can be rapidly performed in a clinical environment to effectively contribute to patient care decisions and public health maneuvers., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)
- Published
- 2014
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19. Natural disruption of two regulatory networks in serotype M3 group A Streptococcus isolates contributes to the virulence factor profile of this hypervirulent serotype.
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Cao TN, Liu Z, Cao TH, Pflughoeft KJ, Treviño J, Danger JL, Beres SB, Musser JM, and Sumby P
- Subjects
- Bacterial Proteins genetics, Gene Deletion, Polymorphism, Single Nucleotide, RNA, Messenger genetics, RNA, Messenger metabolism, Serotyping, Time Factors, Virulence, Virulence Factors genetics, Bacterial Proteins metabolism, Gene Expression Regulation, Bacterial physiology, Streptococcus pyogenes metabolism, Streptococcus pyogenes pathogenicity, Virulence Factors metabolism
- Abstract
Despite the public health challenges associated with the emergence of new pathogenic bacterial strains and/or serotypes, there is a dearth of information regarding the molecular mechanisms that drive this variation. Here, we began to address the mechanisms behind serotype-specific variation between serotype M1 and M3 strains of the human pathogen Streptococcus pyogenes (the group A Streptococcus [GAS]). Spatially diverse contemporary clinical serotype M3 isolates were discovered to contain identical inactivating mutations within genes encoding two regulatory systems that control the expression of important virulence factors, including the thrombolytic agent streptokinase, the protease inhibitor-binding protein-G-related α2-macroglobulin-binding (GRAB) protein, and the antiphagocytic hyaluronic acid capsule. Subsequent analysis of a larger collection of isolates determined that M3 GAS, since at least the 1920s, has harbored a 4-bp deletion in the fasC gene of the fasBCAX regulatory system and an inactivating polymorphism in the rivR regulator-encoding gene. The fasC and rivR mutations in M3 isolates directly affect the virulence factor profile of M3 GAS, as evident by a reduction in streptokinase expression and an enhancement of GRAB expression. Complementation of the fasC mutation in M3 GAS significantly enhanced levels of the small regulatory RNA FasX, which in turn enhanced streptokinase expression. Complementation of the rivR mutation in M3 GAS restored the regulation of grab mRNA abundance but did not alter capsule mRNA levels. While important, the fasC and rivR mutations do not provide a full explanation for why serotype M3 strains are associated with unusually severe invasive infections; thus, further investigation is warranted.
- Published
- 2014
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20. Evolutionary pathway to increased virulence and epidemic group A Streptococcus disease derived from 3,615 genome sequences.
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Nasser W, Beres SB, Olsen RJ, Dean MA, Rice KA, Long SW, Kristinsson KG, Gottfredsson M, Vuopio J, Raisanen K, Caugant DA, Steinbakk M, Low DE, McGeer A, Darenberg J, Henriques-Normark B, Van Beneden CA, Hoffmann S, and Musser JM
- Subjects
- Animals, Base Sequence, Disease Models, Animal, Fasciitis, Necrotizing epidemiology, Fasciitis, Necrotizing genetics, Fasciitis, Necrotizing microbiology, Finland epidemiology, Genes, Bacterial genetics, Genomics, Humans, INDEL Mutation genetics, Pharyngitis epidemiology, Pharyngitis genetics, Pharyngitis microbiology, Polymorphism, Single Nucleotide genetics, Primates microbiology, Selection, Genetic, Serotyping, Streptococcal Infections microbiology, Streptococcus pyogenes isolation & purification, Time Factors, Virulence genetics, Epidemics, Evolution, Molecular, Genome, Bacterial genetics, Streptococcal Infections epidemiology, Streptococcal Infections genetics, Streptococcus pyogenes genetics, Streptococcus pyogenes pathogenicity
- Abstract
We sequenced the genomes of 3,615 strains of serotype Emm protein 1 (M1) group A Streptococcus to unravel the nature and timing of molecular events contributing to the emergence, dissemination, and genetic diversification of an unusually virulent clone that now causes epidemic human infections worldwide. We discovered that the contemporary epidemic clone emerged in stepwise fashion from a precursor cell that first contained the phage encoding an extracellular DNase virulence factor (streptococcal DNase D2, SdaD2) and subsequently acquired the phage encoding the SpeA1 variant of the streptococcal pyrogenic exotoxin A superantigen. The SpeA2 toxin variant evolved from SpeA1 by a single-nucleotide change in the M1 progenitor strain before acquisition by horizontal gene transfer of a large chromosomal region encoding secreted toxins NAD(+)-glycohydrolase and streptolysin O. Acquisition of this 36-kb region in the early 1980s into just one cell containing the phage-encoded sdaD2 and speA2 genes was the final major molecular event preceding the emergence and rapid intercontinental spread of the contemporary epidemic clone. Thus, we resolve a decades-old controversy about the type and sequence of genomic alterations that produced this explosive epidemic. Analysis of comprehensive, population-based contemporary invasive strains from seven countries identified strong patterns of temporal population structure. Compared with a preepidemic reference strain, the contemporary clone is significantly more virulent in nonhuman primate models of pharyngitis and necrotizing fasciitis. A key finding is that the molecular evolutionary events transpiring in just one bacterial cell ultimately have produced millions of human infections worldwide.
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- 2014
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21. Human disease isolates of serotype m4 and m22 group a streptococcus lack genes required for hyaluronic acid capsule biosynthesis.
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Flores AR, Jewell BE, Fittipaldi N, Beres SB, and Musser JM
- Subjects
- Genes, Bacterial, Glucuronosyltransferase deficiency, Humans, Hyaluronan Synthases, Serotyping, Streptococcal Infections microbiology, Streptococcus pyogenes classification, Streptococcus pyogenes genetics, Bacterial Capsules metabolism, Biosynthetic Pathways, Hyaluronic Acid metabolism, Streptococcus pyogenes enzymology, Streptococcus pyogenes metabolism
- Abstract
Unlabelled: Group A streptococcus (GAS) causes human pharyngitis and invasive infections and frequently colonizes individuals asymptomatically. Many lines of evidence generated over decades have shown that the hyaluronic acid capsule is a major virulence factor contributing to these infections. While conducting a whole-genome analysis of the in vivo molecular genetic changes that occur in GAS during longitudinal human pharyngeal interaction, we discovered that serotypes M4 and M22 GAS strains lack the hasABC genes necessary for hyaluronic acid capsule biosynthesis. Using targeted PCR, we found that all 491 temporally and geographically diverse disease isolates of these two serotypes studied lack the hasABC genes. Consistent with the lack of capsule synthesis genes, none of the strains produced detectable hyaluronic acid. Despite the lack of a hyaluronic acid capsule, all strains tested multiplied extensively ex vivo in human blood. Thus, counter to the prevailing concept in GAS pathogenesis research, strains of these two serotypes do not require hyaluronic acid to colonize the upper respiratory tract or cause abundant mucosal or invasive human infections. We speculate that serotype M4 and M22 GAS have alternative, compensatory mechanisms that promote virulence., Importance: A century of study of the antiphagocytic hyaluronic acid capsule made by group A streptococcus has led to the concept that it is a major virulence factor contributing to human pharyngeal and invasive infections. However, the discovery that some strains that cause abundant human infections lack hyaluronic acid biosynthetic genes and fail to produce this capsule provides a new stimulus for research designed to understand the group A streptococcus factors contributing to pharyngeal infection and invasive disease episodes.
- Published
- 2012
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22. Polymorphisms in regulator of protease B (RopB) alter disease phenotype and strain virulence of serotype M3 group A Streptococcus.
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Olsen RJ, Laucirica DR, Watkins ME, Feske ML, Garcia-Bustillos JR, Vu C, Cantu C, Shelburne SA 3rd, Fittipaldi N, Kumaraswami M, Shea PR, Flores AR, Beres SB, Lovgren M, Tyrrell GJ, Efstratiou A, Low DE, Van Beneden CA, and Musser JM
- Subjects
- Animals, Bacterial Proteins metabolism, DNA, Bacterial chemistry, DNA, Bacterial genetics, Disease Models, Animal, Fasciitis, Necrotizing microbiology, Fasciitis, Necrotizing pathology, Humans, Mice, Mutant Proteins genetics, Mutant Proteins metabolism, Pharyngitis pathology, Sequence Analysis, DNA, Streptococcal Infections pathology, Streptococcus pyogenes classification, Virulence, Virulence Factors metabolism, Bacterial Proteins genetics, Pharyngitis microbiology, Polymorphism, Genetic, Streptococcal Infections microbiology, Streptococcus pyogenes genetics, Streptococcus pyogenes isolation & purification, Virulence Factors genetics
- Abstract
Whole-genome sequencing of serotype M3 group A streptococci (GAS) from oropharyngeal and invasive infections in Ontario recently showed that the gene encoding regulator of protease B (RopB) is highly polymorphic in this population. To test the hypothesis that ropB is under diversifying selective pressure among all serotype M3 GAS strains, we sequenced this gene in 1178 strains collected from different infection types, geographic regions, and time periods. The results confirmed our hypothesis and discovered a significant association between mutant ropB alleles, decreased activity of its major regulatory target SpeB, and pharyngitis. Additionally, isoallelic strains with ropB polymorphisms were significantly less virulent in a mouse model of necrotizing fasciitis. These studies provide a model strategy for applying whole-genome sequencing followed by deep single-gene sequencing to generate new insight to the rapid evolution and virulence regulation of human pathogens.
- Published
- 2012
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23. Genomic analysis of emm59 group A Streptococcus invasive strains, United States.
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Fittipaldi N, Olsen RJ, Beres SB, Van Beneden C, and Musser JM
- Subjects
- Antigens, Bacterial genetics, Bacterial Outer Membrane Proteins genetics, Carrier Proteins genetics, INDEL Mutation, Molecular Typing standards, Phylogeny, Polymorphism, Single Nucleotide, Population Surveillance, Reference Standards, Sequence Analysis, DNA, Streptococcus pyogenes classification, Streptococcus pyogenes isolation & purification, United States, Genome, Bacterial, Streptococcal Infections microbiology, Streptococcus pyogenes genetics
- Abstract
Genomic analysis of type emm59 group A Streptococcus invasive strains isolated in the United States discovered higher than anticipated genetic heterogeneity among strains and identified a heretofore unrecognized monoclonal cluster of invasive infections in the San Francisco Bay area. Heightened monitoring for a potential shift in the epidemic behavior of emm59 group A Streptococcus is warranted.
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- 2012
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24. Full-genome dissection of an epidemic of severe invasive disease caused by a hypervirulent, recently emerged clone of group A Streptococcus.
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Fittipaldi N, Beres SB, Olsen RJ, Kapur V, Shea PR, Watkins ME, Cantu CC, Laucirica DR, Jenkins L, Flores AR, Lovgren M, Ardanuy C, Liñares J, Low DE, Tyrrell GJ, and Musser JM
- Subjects
- Animals, Canada epidemiology, DNA, Bacterial genetics, Disease Models, Animal, Epidemics, Fasciitis, Necrotizing microbiology, Fasciitis, Necrotizing pathology, Female, Genome, Bacterial, Humans, Inverted Repeat Sequences genetics, Macaca fascicularis, Male, Mice, Mice, Hairless, Pharyngitis epidemiology, Pharyngitis microbiology, Phylogeny, Saliva microbiology, Sequence Analysis, DNA methods, Skin Diseases, Bacterial microbiology, Skin Diseases, Bacterial pathology, Streptococcal Infections microbiology, Streptococcal Infections transmission, Streptococcus pyogenes classification, Streptococcus pyogenes growth & development, Streptococcus pyogenes pathogenicity, United States epidemiology, Virulence genetics, Streptococcal Infections epidemiology, Streptococcus pyogenes genetics
- Abstract
Group A Streptococcus (GAS) causes an exceptionally broad range of infections in humans, from relatively mild pharyngitis and skin infections to life-threatening necrotizing fasciitis and toxic shock syndrome. An epidemic of severe invasive human infections caused by type emm59 GAS, heretofore an exceedingly rare cause of disease, spread west to east across Canada over a 3-year period (2006 to 2008). By sequencing the genomes of 601 epidemic, historic, and other emm59 organisms, we discovered that a recently emerged, genetically distinct emm59 clone is responsible for the Canadian epidemic. Using near-real-time genome sequencing, we were able to show spread of the Canadian epidemic clone into the United States. The extensive genome data permitted us to identify patterns of geographic dissemination as well as links between emm59 subclonal lineages that cause infections. Mouse and nonhuman primate models of infection demonstrated that the emerged clone is unusually virulent. Transmission of epidemic emm59 strains may have occurred primarily by skin contact, as suggested by an experimental model of skin transmission. In addition, the emm59 strains had a significantly impaired ability to persist in human saliva and to colonize the oropharynx of mice, and seldom caused human pharyngitis. Our study contributes new information to the rapidly emerging field of molecular pathogenomics of bacterial epidemics and illustrates how full-genome data can be used to precisely illuminate the landscape of strain dissemination during a bacterial epidemic., (Copyright © 2012 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)
- Published
- 2012
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25. Group A Streptococcus emm gene types in pharyngeal isolates, Ontario, Canada, 2002-2010.
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Shea PR, Ewbank AL, Gonzalez-Lugo JH, Martagon-Rosado AJ, Martinez-Gutierrez JC, Rehman HA, Serrano-Gonzalez M, Fittipaldi N, Beres SB, Flores AR, Low DE, Willey BM, and Musser JM
- Subjects
- Alleles, Child, Child, Preschool, Genotype, Humans, Infant, Ontario epidemiology, Pharynx microbiology, Phylogeography, Streptococcus pyogenes genetics, Streptococcus pyogenes isolation & purification, Antigens, Bacterial genetics, Bacterial Outer Membrane Proteins genetics, Carrier Proteins genetics, Pharyngitis epidemiology, Pharyngitis microbiology, Streptococcal Infections epidemiology, Streptococcus pyogenes classification
- Abstract
Group A Streptococcus (GAS) is a human-adapted pathogen that causes a variety of diseases, including pharyngitis and invasive infections. GAS strains are categorized by variation in the nucleotide sequence of the gene (emm) that encodes the M protein. To identify the emm types of GAS strains causing pharyngitis in Ontario, Canada, we sequenced the hypervariable region of the emm gene in 4,635 pharyngeal GAS isolates collected during 2002-2010. The most prevalent emm types varied little from year to year. In contrast, fine-scale geographic analysis identified inter-site variability in the most common emm types. Additionally, we observed fluctuations in yearly frequency of emm3 strains from pharyngitis patients that coincided with peaks of emm3 invasive infections. We also discovered a striking increase in frequency of emm89 strains among isolates from patients with pharyngitis and invasive disease. These findings about the epidemiology of GAS are potentially useful for vaccine research.
- Published
- 2011
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26. Evolution of diversity in epidemics revealed by analysis of the human bacterial pathogen group A Streptococcus.
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Carroll RK, Beres SB, Sitkiewicz I, Peterson L, Matsunami RK, Engler DA, Flores AR, Sumby P, and Musser JM
- Subjects
- Biodiversity, Biological Evolution, Gene Expression Regulation, Genome, Bacterial genetics, Humans, Microarray Analysis, Phenotype, Proteomics, Streptococcal Infections microbiology, Streptococcus pyogenes classification, Streptococcus pyogenes metabolism, Streptococcus pyogenes pathogenicity, Virulence Factors genetics, Bacterial Proteins genetics, Exotoxins genetics, Polymorphism, Genetic, Streptococcal Infections epidemiology, Streptococcal Infections genetics, Streptococcus pyogenes genetics
- Abstract
Advancements in high-throughput, high-volume data generating techniques increasingly present us with opportunities to probe new areas of biology. In this work we assessed the extent to which four closely related and genetically representative strains of group A Streptococcus causing epidemic disease have differentiated from one another. Comparative genome sequencing, expression microarray analysis, and proteomic studies were used in parallel to assess strain variation. The extent of phenotypic differentiation was unexpectedly large. We found significant associations between genetic polymorphisms and alterations in gene expression allowing us to estimate the frequency with which specific types of polymorphisms alter gene transcription. We identified polymorphisms in the gene (ropB) encoding the RopB regulator that associate with altered transcription of speB and production of the SpeB protein, a critical secreted protease virulence factor. Although these four epidemic strains are closely related, a key discovery is that accumulation of modest genetic changes has rapidly resulted in significant strain phenotypic differentiation, including the extracellular proteome that contains multiple virulence factors. These data provide enhanced understanding of genetic events resulting in strain variation in bacterial epidemics., (Copyright © 2011 Elsevier B.V. All rights reserved.)
- Published
- 2011
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27. Distinct signatures of diversifying selection revealed by genome analysis of respiratory tract and invasive bacterial populations.
- Author
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Shea PR, Beres SB, Flores AR, Ewbank AL, Gonzalez-Lugo JH, Martagon-Rosado AJ, Martinez-Gutierrez JC, Rehman HA, Serrano-Gonzalez M, Fittipaldi N, Ayers SD, Webb P, Willey BM, Low DE, and Musser JM
- Subjects
- Animals, Female, Genome-Wide Association Study, Humans, Male, Primates, Evolution, Molecular, Genome, Bacterial physiology, Pharyngitis genetics, Phylogeny, Streptococcal Infections genetics, Streptococcus pyogenes genetics
- Abstract
Many pathogens colonize different anatomical sites, but the selective pressures contributing to survival in the diverse niches are poorly understood. Group A Streptococcus (GAS) is a human-adapted bacterium that causes a range of infections. Much effort has been expended to dissect the molecular basis of invasive (sterile-site) infections, but little is known about the genomes of strains causing pharyngitis (streptococcal "sore throat"). Additionally, there is essentially nothing known about the genetic relationships between populations of invasive and pharyngitis strains. In particular, it is unclear if invasive strains represent a distinct genetic subpopulation of strains that cause pharyngitis. We compared the genomes of 86 serotype M3 GAS pharyngitis strains with those of 215 invasive M3 strains from the same geographical location. The pharyngitis and invasive groups were highly related to each other and had virtually identical phylogenetic structures, indicating they belong to the same genetic pool. Despite the overall high degree of genetic similarity, we discovered that strains from different host environments (i.e., throat, normally sterile sites) have distinct patterns of diversifying selection at the nucleotide level. In particular, the pattern of polymorphisms in the hyaluronic acid capsule synthesis operon was especially different between the two strain populations. This finding was mirrored by data obtained from full-genome analysis of strains sequentially cultured from nonhuman primates. Our results answer the long-standing question of the genetic relationship between GAS pharyngitis and invasive strains. The data provide previously undescribed information about the evolutionary history of pathogenic microbes that cause disease in different anatomical sites.
- Published
- 2011
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28. Virulence of serotype M3 Group A Streptococcus strains in wax worms (Galleria mellonella larvae).
- Author
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Olsen RJ, Watkins ME, Cantu CC, Beres SB, and Musser JM
- Subjects
- Animals, Antigens, Bacterial genetics, Bacterial Outer Membrane Proteins genetics, Carrier Proteins genetics, Carrier State microbiology, Genotype, Humans, Larva microbiology, Lethal Dose 50, Serotyping, Streptococcal Infections microbiology, Streptococcus pyogenes classification, Streptococcus pyogenes isolation & purification, Survival Analysis, Virulence, Host-Pathogen Interactions, Lepidoptera microbiology, Streptococcus pyogenes pathogenicity
- Abstract
Group A Streptococcus (GAS) causes human infections that range in severity from pharyngitis ("strep-throat") to necrotizing fasciitis ("flesh-eating disease"). To facilitate investigation of the molecular basis of host-pathogen interactions, infection models capable of rapidly screening for differences in GAS strain virulence are needed. To this end, we developed a Galleria mellonella larvae (wax worm) model of invasive GAS infection and used it to compare the virulence of serotype M3 GAS strains. We found that GAS causes severe tissue damage and kills wax worms in a dose-dependent manner. The virulence of genetically distinct GAS strains was compared by Kaplan-Meier survival analysis and determining 50% lethal doses (LD 50). Host-pathogen interactions were further characterized using quantitative culture, histopathology and TaqMan assays. GAS strains known to be highly pathogenic in mice and monkeys caused significantly lower survival and had significantly lower LD 50s in wax worms than GAS strains associated with attenuated virulence or asymptomatic carriage. Furthermore, isogenic inactivation of proven virulence factors resulted in a significantly increased LD 50 and decreased lesion size compared to the wild-type strain, a finding that also strongly correlates with animal studies. Importantly, survival analysis and LD 50 determination in wax worms supported our hypothesis that a newly emerged GAS subclone that is epidemiologically associated with more human necrotizing fasciitis cases than its progenitor lineage has significantly increased virulence. We conclude that GAS virulence in wax worms strongly correlates with the data obtained in vertebrate models, and thus, the Galleria mellonella larva is a useful host organism to study GAS pathogenesis.
- Published
- 2011
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29. Molecular complexity of successive bacterial epidemics deconvoluted by comparative pathogenomics.
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Beres SB, Carroll RK, Shea PR, Sitkiewicz I, Martinez-Gutierrez JC, Low DE, McGeer A, Willey BM, Green K, Tyrrell GJ, Goldman TD, Feldgarden M, Birren BW, Fofanov Y, Boos J, Wheaton WD, Honisch C, and Musser JM
- Subjects
- Biological Evolution, Codon, Terminator, Genotype, Humans, Mass Spectrometry, Oligonucleotide Array Sequence Analysis, Ontario epidemiology, Phenotype, Phylogeny, Polymorphism, Single Nucleotide, Streptococcus pyogenes pathogenicity, Virulence, Disease Outbreaks, Genome, Bacterial, Streptococcal Infections epidemiology, Streptococcus pyogenes isolation & purification
- Abstract
Understanding the fine-structure molecular architecture of bacterial epidemics has been a long-sought goal of infectious disease research. We used short-read-length DNA sequencing coupled with mass spectroscopy analysis of SNPs to study the molecular pathogenomics of three successive epidemics of invasive infections involving 344 serotype M3 group A Streptococcus in Ontario, Canada. Sequencing the genome of 95 strains from the three epidemics, coupled with analysis of 280 biallelic SNPs in all 344 strains, revealed an unexpectedly complex population structure composed of a dynamic mixture of distinct clonally related complexes. We discovered that each epidemic is dominated by micro- and macrobursts of multiple emergent clones, some with distinct strain genotype-patient phenotype relationships. On average, strains were differentiated from one another by only 49 SNPs and 11 insertion-deletion events (indels) in the core genome. Ten percent of SNPs are strain specific; that is, each strain has a unique genome sequence. We identified nonrandom temporal-spatial patterns of strain distribution within and between the epidemic peaks. The extensive full-genome data permitted us to identify genes with significantly increased rates of nonsynonymous (amino acid-altering) nucleotide polymorphisms, thereby providing clues about selective forces operative in the host. Comparative expression microarray analysis revealed that closely related strains differentiated by seemingly modest genetic changes can have significantly divergent transcriptomes. We conclude that enhanced understanding of bacterial epidemics requires a deep-sequencing, geographically centric, comparative pathogenomics strategy.
- Published
- 2010
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30. Contribution of AmyA, an extracellular alpha-glucan degrading enzyme, to group A streptococcal host-pathogen interaction.
- Author
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Shelburne Iii SA, Keith DB, Davenport MT, Beres SB, Carroll RK, and Musser JM
- Subjects
- Animals, Bacterial Proteins genetics, Cell Line, Cyclodextrins biosynthesis, Female, Genetic Complementation Test, Glucans metabolism, Glucosyltransferases genetics, Humans, Mice, Mutagenesis, Insertional, Mutation, Pharyngitis microbiology, RNA, Bacterial genetics, Streptococcal Infections microbiology, Streptococcus pyogenes genetics, Streptococcus pyogenes pathogenicity, Virulence, Bacterial Proteins metabolism, Glucosyltransferases metabolism, Host-Pathogen Interactions, Streptococcus pyogenes enzymology
- Abstract
alpha-Glucans such as starch and glycogen are abundant in the human oropharynx, the main site of group A Streptococcus (GAS) infection. However, the role in pathogenesis of GAS extracellular alpha-glucan binding and degrading enzymes is unknown. The serotype M1 GAS genome encodes two extracellular proteins putatively involved in alpha-glucan binding and degradation; pulA encodes a cell wall anchored pullulanase and amyA encodes a freely secreted putative cyclomaltodextrin alpha-glucanotransferase. Genetic inactivation of amyA, but not pulA, abolished GAS alpha-glucan degradation. The DeltaamyA strain had a slower rate of translocation across human pharyngeal epithelial cells. Consistent with this finding, the DeltaamyA strain was less virulent following mouse mucosal challenge. Recombinant AmyA degraded alpha-glucans into beta-cyclomaltodextrins that reduced pharyngeal cell transepithelial resistance, providing a physiologic explanation for the observed transepithelial migration phenotype. Higher amyA transcript levels were present in serotype M1 GAS strains causing invasive infection compared with strains causing pharyngitis. GAS proliferation in a defined alpha-glucan-containing medium was dependent on the presence of human salivary alpha-amylase. These data delineate the molecular mechanisms by which alpha-glucan degradation contributes to GAS host-pathogen interaction, including how GAS uses human salivary alpha-amylase for its own metabolic benefit.
- Published
- 2009
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31. Contribution of exogenous genetic elements to the group A Streptococcus metagenome.
- Author
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Beres SB and Musser JM
- Subjects
- Bacterial Proteins genetics, Bacterial Proteins metabolism, Drug Resistance, Microbial genetics, Humans, Metagenomics, Models, Genetic, Tetracycline pharmacology, Genome, Bacterial, Streptococcus pyogenes genetics
- Abstract
Variation in gene content among strains of a bacterial species contributes to biomedically relevant differences in phenotypes such as virulence and antimicrobial resistance. Group A Streptococcus (GAS) causes a diverse array of human infections and sequelae, and exhibits a complex pathogenic behavior. To enhance our understanding of genotype-phenotype relationships in this important pathogen, we determined the complete genome sequences of four GAS strains expressing M protein serotypes (M2, M4, and 2 M12) that commonly cause noninvasive and invasive infections. These sequences were compared with eight previously determined GAS genomes and regions of variably present gene content were assessed. Consistent with the previously determined genomes, each of the new genomes is approximately 1.9 Mb in size, with approximately 10% of the gene content of each encoded on variably present exogenous genetic elements. Like the other GAS genomes, these four genomes are polylysogenic and prophage encode the majority of the variably present gene content of each. In contrast to most of the previously determined genomes, multiple exogenous integrated conjugative elements (ICEs) with characteristics of conjugative transposons and plasmids are present in these new genomes. Cumulatively, 242 new GAS metagenome genes were identified that were not present in the previously sequenced genomes. Importantly, ICEs accounted for 41% of the new GAS metagenome gene content identified in these four genomes. Two large ICEs, designated 2096-RD.2 (63 kb) and 10750-RD.2 (49 kb), have multiple genes encoding resistance to antimicrobial agents, including tetracycline and erythromycin, respectively. Also resident on these ICEs are three genes encoding inferred extracellular proteins of unknown function, including a predicted cell surface protein that is only present in the genome of the serotype M12 strain cultured from a patient with acute poststreptococcal glomerulonephritis. The data provide new information about the GAS metagenome and will assist studies of pathogenesis, antimicrobial resistance, and population genomics.
- Published
- 2007
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32. Molecular genetic anatomy of inter- and intraserotype variation in the human bacterial pathogen group A Streptococcus.
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Beres SB, Richter EW, Nagiec MJ, Sumby P, Porcella SF, DeLeo FR, and Musser JM
- Subjects
- Amino Acid Sequence, Animals, Base Sequence, Gene Expression Profiling, Genes, Bacterial, Genome, Bacterial, Genotype, Humans, Mice, Molecular Biology, Molecular Sequence Data, Oligonucleotide Array Sequence Analysis, Phenotype, Phylogeny, Sequence Alignment, Serotyping, Streptococcus pyogenes pathogenicity, Genetic Variation, Sequence Analysis, DNA, Streptococcal Infections, Streptococcus pyogenes genetics
- Abstract
In recent years we have studied the relationship between strain genotypes and patient phenotypes in group A Streptococcus (GAS), a model human bacterial pathogen that causes extensive morbidity and mortality worldwide. We have concentrated our efforts on serotype M3 organisms because these strains are common causes of pharyngeal and invasive infections, produce unusually severe invasive infections, and can exhibit epidemic behavior. Our studies have been hindered by the lack of genome-scale phylogenies of multiple GAS strains and whole-genome sequences of multiple serotype M3 strains recovered from individuals with defined clinical phenotypes. To remove some of these impediments, we sequenced to closure the genome of four additional GAS strains and conducted comparative genomic resequencing of 12 contemporary serotype M3 strains representing distinct genotypes and phenotypes. Serotype M3 strains are a single phylogenetic lineage. Strains from asymptomatic throat carriers were significantly less virulent for mice than sterile-site isolates and evolved to a less virulent phenotype by multiple genetic pathways. Strain persistence or extinction between epidemics was strongly associated with presence or absence, respectively, of the prophage encoding streptococcal pyrogenic exotoxin A. A serotype M3 clone significantly underrepresented among necrotizing fasciitis cases has a unique frameshift mutation that truncates MtsR, a transcriptional regulator controlling expression of genes encoding iron-acquisition proteins. Expression microarray analysis of this clone confirmed significant alteration in expression of genes encoding iron metabolism proteins. Our analysis provided unprecedented detail about the molecular anatomy of bacterial strain genotype-patient phenotype relationships.
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- 2006
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33. Genome sequence of a serotype M28 strain of group a streptococcus: potential new insights into puerperal sepsis and bacterial disease specificity.
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Green NM, Zhang S, Porcella SF, Nagiec MJ, Barbian KD, Beres SB, LeFebvre RB, and Musser JM
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- Antigens, Bacterial genetics, Bacterial Outer Membrane Proteins genetics, Base Sequence, Blotting, Western, Carrier Proteins genetics, DNA, Bacterial chemistry, DNA, Bacterial genetics, Female, Humans, Phylogeny, Polymerase Chain Reaction, Prophages genetics, Recombinant Proteins, Sepsis microbiology, Sequence Alignment, Streptococcus pyogenes pathogenicity, Virulence Factors, Genome, Bacterial, Puerperal Infection microbiology, Streptococcal Infections microbiology, Streptococcus pyogenes genetics
- Abstract
Puerperal sepsis, a major cause of death of young women in Europe in the 1800s, was due predominantly to the gram-positive pathogen group A Streptococcus. Studies conducted during past decades have shown that serotype M28 strains are the major group A Streptococcus organisms responsible for many of these infections. To begin to increase our understanding of their enrichment in puerperal sepsis, we sequenced the genome of a genetically representative strain. This strain has genes encoding a novel array of prophage virulence factors, cell-surface proteins, and other molecules likely to contribute to host-pathogen interactions. Importantly, genes for 7 inferred extracellular proteins are encoded by a 37.4-kb foreign DNA element that is shared with group B Streptococcus and is present in all serotype M28 strains. Proteins encoded by the 37.4-kb element were expressed extracellularly and in human infections. Acquisition of foreign genes has helped create a disease-specialist clone of this pathogen.
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- 2005
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34. Genetic diversity among type emm28 group A Streptococcus strains causing invasive infections and pharyngitis.
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Green NM, Beres SB, Graviss EA, Allison JE, McGeer AJ, Vuopio-Varkila J, LeFebvre RB, and Musser JM
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- Chromosome Mapping, Drug Resistance, Bacterial, Genetic Variation, Humans, Macrolides pharmacology, Prophages genetics, Streptococcus pyogenes pathogenicity, Virulence genetics, Antigens, Bacterial genetics, Bacterial Outer Membrane Proteins genetics, Carrier Proteins genetics, Pharyngitis microbiology, Streptococcal Infections microbiology, Streptococcus pyogenes genetics
- Abstract
Genome sequencing of group A Streptococcus (GAS) has revealed that prophages account for the vast majority of gene content differences between strains. Serotype M28 strains are a leading cause of pharyngitis and invasive infections, but little is known about genetic diversity present in natural populations of these organisms. To study this issue, population-based samples of 568 strains from Ontario, Canada; Finland; and Houston, Texas, were analyzed. Special attention was given to analysis of variation in prophage-encoded virulence gene content by a PCR-based method. Thirty and 29 distinct prophage-encoded virulence gene profiles were identified among pharyngitis and invasive infection isolates. Thirteen profiles, representing the majority of the strains, were shared between these two classes of isolates. Significant differences were observed in the frequency of occurrence of certain prophage toxin gene profiles and infection type. M28 strains are highly diverse in prophage-encoded virulence gene content and integration site, supporting the key concept that prophages are critical contributors to GAS genetic diversity and population biology. Nucleotide sequence variation in the emm gene (encodes M protein) was also examined. Only three allelic variants were identified in the hypervariable portion of the emm28 gene. All but one strain had the same inferred amino acid sequence in the first 100 amino acids of the mature M28 protein. In contrast, size differences in the emm28 gene and inferred protein due to variable numbers of C-terminal repeats were common. The presence of macrolide resistance genes (mefA, ermB, and ermTR) was analyzed by PCR, and less than 2% of the strains were positive.
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- 2005
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35. Analysis of a novel prophage-encoded group A Streptococcus extracellular phospholipase A(2).
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Nagiec MJ, Lei B, Parker SK, Vasil ML, Matsumoto M, Ireland RM, Beres SB, Hoe NP, and Musser JM
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- Alleles, Amino Acid Sequence, Base Sequence, Humans, Molecular Sequence Data, Phospholipases A genetics, Phospholipases A physiology, Prophages genetics, Structure-Activity Relationship, Substrate Specificity, Phospholipases A chemistry, Streptococcus pyogenes enzymology
- Abstract
Group A Streptococcus (GAS) is an important human pathogen that causes many types of infections, including pharyngitis and severe invasive diseases. We recently sequenced the genome of a serotype M3 strain and identified a prophage-encoded secreted phospholipase A(2) designated SlaA. To study SlaA structure-activity relationships, 20 site-specific mutants were constructed by alanine-replacement mutagenesis and purified to apparent homogeneity. Enzymatic activity was greatly reduced by alanine replacement of amino acid residues previously described as crucial in the catalytic mechanism of secreted phospholipase A(2). Similarly, substitution of five residues in an inferred Ca(2+)-binding loop and three residues in the inferred active site region resulted in loss of activity of 76.5% or greater relative to the wild-type enzyme. Analysis of enzyme substrate specificity confirmed SlaA as a phospholipase A(2), with activity against multiple phospholipid head groups and acyl chains located at the sn-2 position. PCR analysis of 1,189 GAS strains representing 48 M protein serotypes commonly causing human infections identified the slaA gene in 129 strains of nine serotypes (M1, M2, M3, M4, M6, M22, M28, M75, and st3757). Expression of SlaA by strains of these serotypes was confirmed by Western immunoblot. SlaA production increased rapidly and substantially on co-culture with Detroit 562 human pharyngeal epithelial cells. Together, these data provide new information about a novel extracellular enzyme that participates in GAS-human interactions.
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- 2004
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36. Progress toward characterization of the group A Streptococcus metagenome: complete genome sequence of a macrolide-resistant serotype M6 strain.
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Banks DJ, Porcella SF, Barbian KD, Beres SB, Philips LE, Voyich JM, DeLeo FR, Martin JM, Somerville GA, and Musser JM
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- Alleles, Amino Acid Motifs genetics, Antibodies, Bacterial blood, Bacterial Outer Membrane Proteins genetics, Bacterial Proteins genetics, Bacterial Proteins immunology, Blotting, Western, Child, Drug Resistance, Bacterial genetics, Exotoxins genetics, Flow Cytometry, Humans, Membrane Proteins genetics, Membrane Proteins immunology, Pharyngitis blood, Pharyngitis microbiology, Streptococcal Infections blood, Streptococcal Infections microbiology, Streptococcus Phages genetics, Streptococcus pyogenes drug effects, Streptococcus pyogenes isolation & purification, Genome, Bacterial, Macrolides pharmacology, R Factors genetics, Streptococcus pyogenes genetics
- Abstract
We describe the genome sequence of a macrolide-resistant strain (MGAS10394) of serotype M6 group A Streptococcus (GAS). The genome is 1,900,156 bp in length, and 8 prophage-like elements or remnants compose 12.4% of the chromosome. A 8.3-kb prophage remnant encodes the SpeA4 variant of streptococcal pyrogenic exotoxin A. The genome of strain MGAS10394 contains a chimeric genetic element composed of prophage genes and a transposon encoding the mefA gene conferring macrolide resistance. This chimeric element also has a gene encoding a novel surface-exposed protein (designated "R6 protein"), with an LPKTG cell-anchor motif located at the carboxyterminus. Surface expression of this protein was confirmed by flow cytometry. Humans with GAS pharyngitis caused by serotype M6 strains had antibody against the R6 protein present in convalescent, but not acute, serum samples. Our studies add to the theme that GAS prophage-encoded extracellular proteins contribute to host-pathogen interactions in a strain-specific fashion.
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- 2004
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37. Genome-wide molecular dissection of serotype M3 group A Streptococcus strains causing two epidemics of invasive infections.
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Beres SB, Sylva GL, Sturdevant DE, Granville CN, Liu M, Ricklefs SM, Whitney AR, Parkins LD, Hoe NP, Adams GJ, Low DE, DeLeo FR, McGeer A, and Musser JM
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- Antigens, Bacterial genetics, Antigens, Bacterial physiology, Bacterial Outer Membrane Proteins genetics, Bacterial Outer Membrane Proteins physiology, Bacterial Proteins genetics, Canada epidemiology, Carrier Proteins genetics, Carrier Proteins physiology, Chromosome Aberrations, Genetic Variation, Genomics methods, Genotype, Humans, Polymorphism, Single Nucleotide, Population Surveillance, Sequence Analysis, Streptococcal Infections virology, Streptococcus pyogenes immunology, Streptococcus pyogenes pathogenicity, Virulence genetics, Virulence Factors genetics, Disease Outbreaks, Genome, Bacterial, Streptococcal Infections epidemiology, Streptococcus pyogenes genetics
- Abstract
Molecular factors that contribute to the emergence of new virulent bacterial subclones and epidemics are poorly understood. We hypothesized that analysis of a population-based strain sample of serotype M3 group A Streptococcus (GAS) recovered from patients with invasive infection by using genome-wide investigative methods would provide new insight into this fundamental infectious disease problem. Serotype M3 GAS strains (n = 255) cultured from patients in Ontario, Canada, over 11 years and representing two distinct infection peaks were studied. Genetic diversity was indexed by pulsed-field gel electrophoresis, DNA-DNA microarray, whole-genome PCR scanning, prophage genotyping, targeted gene sequencing, and single-nucleotide polymorphism genotyping. All variation in gene content was attributable to acquisition or loss of prophages, a molecular process that generated unique combinations of proven or putative virulence genes. Distinct serotype M3 genotypes experienced rapid population expansion and caused infections that differed significantly in character and severity. Molecular genetic analysis, combined with immunologic studies, implicated a 4-aa duplication in the extreme N terminus of M protein as a factor contributing to an epidemic wave of serotype M3 invasive infections. This finding has implications for GAS vaccine research. Genome-wide analysis of population-based strain samples cultured from clinically well defined patients is crucial for understanding the molecular events underlying bacterial epidemics.
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- 2004
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38. Intrahost sequence variation in the streptococcal inhibitor of complement gene in patients with human pharyngitis.
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Matsumoto M, Hoe NP, Liu M, Beres SB, Sylva GL, Brandt CM, Haase G, and Musser JM
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- Acute Disease, Amino Acid Sequence, Base Sequence, Cloning, Molecular, Collagen genetics, Gene Deletion, Genetic Variation, Humans, Microsatellite Repeats, Molecular Sequence Data, Pharynx microbiology, Streptococcus pyogenes chemistry, Bacterial Proteins genetics, Pharyngitis microbiology, Streptococcal Infections microbiology, Streptococcus pyogenes genetics
- Abstract
Selection of new variants of the streptococcal inhibitor of complement protein has been implicated in the perpetuation of epidemics caused by serotype M1 strains of group A Streptococcus (GAS). The frequency at which new streptococcal inhibitor of complement (Sic) variants arise in an infected individual is not known. To study this issue, the sic gene was sequenced in 100 isolates cultured from throat swabs of each of 20 patients with acute pharyngitis caused by serotype M1 GAS. Five patients were infected with GAS populations expressing 2 Sic variants characterized by deletion of a region of the protein. In contrast, no intrahost variation was detected in the number of a pentanucleotide repeat (CAAAA) that controls production of a bacterial cell-surface collagen-like protein by slipped-strand mispairing. Sic variation occurs at a sufficient frequency in vivo to result in mixed infections on the mucosal surface of human hosts, potentially contributing to pathogen survival.
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- 2003
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39. The fundamental contribution of phages to GAS evolution, genome diversification and strain emergence.
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Banks DJ, Beres SB, and Musser JM
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- Amino Acid Sequence, Anti-Bacterial Agents chemistry, Drug Resistance genetics, Gene Expression Regulation, Viral, Lysogeny, Macrolides, Streptococcus Phages immunology, Virulence Factors immunology, Evolution, Molecular, Genome, Bacterial, Streptococcus Phages genetics, Streptococcus pyogenes genetics, Streptococcus pyogenes virology
- Abstract
The human bacterial pathogen group A Streptococcus (GAS) causes many different diseases including pharyngitis, tonsillitis, impetigo, scarlet fever, streptococcal toxic shock syndrome, necrotizing fasciitis and myositis, and the post-infection sequelae glomerulonephritis and rheumatic fever. The frequency and severity of GAS infections increased in the 1980s and 1990s, but the cause of this increase is unknown. Recently, genome sequencing of serotype M1, M3 and M18 strains revealed many new proven or putative virulence factors that are encoded by phages or phage-like elements. Importantly, these genetic elements account for an unexpectedly large proportion of the difference in gene content between the three strains. These new genome-sequencing studies have provided evidence that temporally and geographically distinct epidemics, and the complex array of GAS clinical presentations, might be related in part to the acquisition or evolution of phage-encoded virulence factors. We anticipate that new phage-encoded virulence factors will be identified by sequencing the genomes of additional GAS strains, including organisms non-randomly associated with particular clinical syndromes.
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- 2002
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40. Genome sequence and comparative microarray analysis of serotype M18 group A Streptococcus strains associated with acute rheumatic fever outbreaks.
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Smoot JC, Barbian KD, Van Gompel JJ, Smoot LM, Chaussee MS, Sylva GL, Sturdevant DE, Ricklefs SM, Porcella SF, Parkins LD, Beres SB, Campbell DS, Smith TM, Zhang Q, Kapur V, Daly JA, Veasy LG, and Musser JM
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- Acute Disease, Base Sequence, Disease Outbreaks, Genetic Variation, Humans, Molecular Sequence Data, Rheumatic Fever etiology, Serotyping, Streptococcus pyogenes classification, Virulence genetics, Genome, Bacterial, Oligonucleotide Array Sequence Analysis, Rheumatic Fever microbiology, Streptococcus pyogenes genetics
- Abstract
Acute rheumatic fever (ARF), a sequelae of group A Streptococcus (GAS) infection, is the most common cause of preventable childhood heart disease worldwide. The molecular basis of ARF and the subsequent rheumatic heart disease are poorly understood. Serotype M18 GAS strains have been associated for decades with ARF outbreaks in the U.S. As a first step toward gaining new insight into ARF pathogenesis, we sequenced the genome of strain MGAS8232, a serotype M18 organism isolated from a patient with ARF. The genome is a circular chromosome of 1,895,017 bp, and it shares 1.7 Mb of closely related genetic material with strain SF370 (a sequenced serotype M1 strain). Strain MGAS8232 has 178 ORFs absent in SF370. Phages, phage-like elements, and insertion sequences are the major sources of variation between the genomes. The genomes of strain MGAS8232 and SF370 encode many of the same proven or putative virulence factors. Importantly, strain MGAS8232 has genes encoding many additional secreted proteins involved in human-GAS interactions, including streptococcal pyrogenic exotoxin A (scarlet fever toxin) and two uncharacterized pyrogenic exotoxin homologues, all phage-associated. DNA microarray analysis of 36 serotype M18 strains from diverse localities showed that most regions of variation were phages or phage-like elements. Two epidemics of ARF occurring 12 years apart in Salt Lake City, UT, were caused by serotype M18 strains that were genetically identical, or nearly so. Our analysis provides a critical foundation for accelerated research into ARF pathogenesis and a molecular framework to study the plasticity of GAS genomes.
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- 2002
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41. A Chimeric Penicillin Binding Protein 2X Significantly Decreases in Vitro Beta-Lactam Susceptibility and Increases in Vivo Fitness of Streptococcus pyogenes
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Olsen, Randall J, Zhu, Luchang, Mangham, Regan E, Faili, Ahmad, Kayal, Samer, Beres, Stephen B, Musser, Andrew, Jonchère, Laurent, Houston Methodist Hospital [Houston, TX, USA], Centre d'Investigation Clinique [Rennes] (CIC), Université de Rennes (UR)-Hôpital Pontchaillou-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Pontchaillou [Rennes], and None
- Subjects
Myositis ,Streptococcus pyogenes ,Recombinant Fusion Proteins ,[SDV]Life Sciences [q-bio] ,Penicillin G ,Penicillins ,beta-Lactams ,Anti-Bacterial Agents ,Pathology and Forensic Medicine ,[SDV] Life Sciences [q-bio] ,Mice ,Streptococcus pneumoniae ,[SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology ,Animals ,Penicillin-Binding Proteins ,Fasciitis, Necrotizing ,[SDV.MP] Life Sciences [q-bio]/Microbiology and Parasitology - Abstract
International audience; All tested strains of Streptococcus pyogenes (group A streptococcus, GAS) remain susceptible to penicillin. However, GAS strains with amino acid substitutions in penicillin-binding proteins that confer decreased susceptibility to beta-lactam antibiotics have been identified recently. This discovery raises concerns about emergence of beta-lactam antibiotic resistance in GAS. Whole genome sequencing recently identified GAS strains with a chimeric penicillin-binding protein 2X (PBP2X) containing a recombinant segment from Streptococcus dysgalactiae subspecies equisimilis (SDSE). To directly test the hypothesis that the chimeric SDSE-like PBP2X alters beta-lactam susceptibility in vitro and fitness in vivo, an isogenic mutant strain was generated and virulence assessed in a mouse model of necrotizing myositis. Compared with naturally occurring and isogenic strains with a wild-type GAS-like PBP2X, strains with the chimeric SDSE-like PBP2X had reduced susceptibility in vitro to nine beta-lactam antibiotics. In a mouse model of necrotizing myositis, the strains had identical fitness in the absence of benzylpenicillin treatment. However, mice treated intermittently with a subtherapeutic dose of benzylpenicillin had significantly more colony-forming units recovered from limbs infected with strains with the chimeric SDSE-like PBP2X. These results show that mutations such as the PBP2X chimera may result in significantly decreased beta-lactam susceptibility and increased fitness and virulence. Expanded diagnostic laboratory surveillance, genome sequencing, and molecular pathogenesis study of potentially emergent beta-lactam antibiotic resistance among GAS are needed.
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- 2022
42. Genetic evolution of invasive emm28 Streptococcus pyogenes strains and significant association with puerperal infections in young women in Finland
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Gröndahl-Yli-Hannuksela, Kirsi, Beres, Stephen B., Hyyryläinen, H.-L., Kallonen, Teemu, Musser, James M., and Vuopio, Jaana
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Adult ,Aged, 80 and over ,Whole-genome sequencing ,Antigens, Bacterial ,Adolescent ,Streptococcus pyogenes ,Group A streptococcus ,Infant, Newborn ,Infant ,Bacteremia ,Middle Aged ,Article ,Young Adult ,Pregnancy ,Child, Preschool ,Streptococcal Infections ,Humans ,Puerperal Infection ,Female ,Puerperal sepsis ,Child ,Bacteraemic ,Finland ,Aged ,Retrospective Studies - Abstract
Objectives Streptococcus pyogenes or group A streptococcus (GAS) is a human specific pathogen that annually infects over 700 million individuals. GAS strains of type emm28 are an abundant cause of invasive infections in Europe and North America. Methods We conducted a population-based study on bacteraemic emm28 GAS cases in Finland, from 1995 to 2015. Whole-genome sequencing (WGS) was used to genetically characterize the bacterial isolates. Bayesian analysis of the population structure was used to define genetic clades. Register-linkage analysis was performed to test for association of emm28 GAS with delivery- or postpartum-related infections. A genome-wide association study was used to search for DNA sequences associated with delivery or puerperal infections. Results Among 3060 bacteraemic cases reported during the study period, 714 were caused by emm28. Women comprised a majority of cases (59 %, 422/714), and were significantly over-represented (84.4 %, 162/192, p < 0.0001) among cases in the childbearing age group (20–40 years). Register-linkage analysis revealed strong association (p < 0.0001) of emm28 bacteraemias with delivery and puerperium. In this register-linkage analysis, 120 women with GAS bacteraemia were identified and linked to delivery, infections during delivery or puerperium time. Among these the proportion of cases caused by emm28 was significantly higher than any other emm type (55.8%, 67/120, p < 0.0001). Among the four genetic subclades identified, SC1B has dominated among the bacteraemic cases since 2000. Altogether 620 of 653 (94.9%) isolates belonged to SC1B. No specific sequence or genetic clade was found nonrandomly associated with delivery or puerperal infections. Conclusions Women of childbearing age were significantly overrepresented among bacteraemic emm28 GAS cases, and in particular were strongly associated with delivery and puerperium cases over the 21 years studied. The molecular mechanisms behind these associations are unclear and warrant further investigation.
- Published
- 2020
43. Population Genomic Molecular Epidemiological Study of Macrolide-Resistant Streptococcus pyogenes in Iceland, 1995 to 2016: Identification of a Large Clonal Population with a pbp2x Mutation Conferring Reduced In Vitro β-Lactam Susceptibility
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Southon, Sara B., Beres, Stephen B., Kachroo, Priyanka, Saavedra, Matthew Ojeda, Erlendsdóttir, Helga, Haraldsson, Gunnsteinn, Yerramilli, Prasanti, Pruitt, Layne, Zhu, Luchang, Musser, James M., Kristinsson, Karl G., Læknadeild (HÍ), Faculty of Medicine (UI), Heilbrigðisvísindasvið (HÍ), School of Health Sciences (UI), Háskóli Íslands, and University of Iceland
- Subjects
Antibiotic resistance ,Streptococcus pyogenes ,Molecular epidemiology ,Sýkingar ,Macrolides ,Population genomics ,Erfðarannsóknir ,Sýklalyf - Abstract
Publisher's version (útgefin grein), Resistance to macrolide antibiotics is a global concern in the treatment of Streptococcus pyogenes (group A Streptococcus [GAS]) infections. In Iceland, since the detection of the first macrolide-resistant isolate in 1998, three epidemic waves of macrolide-resistant GAS infections have occurred, with peaks in 1999, 2004, and 2008. We conducted whole-genome sequencing of all 1,575 available GAS macrolide-resistant clinical isolates of all infection types collected at the national reference laboratory in Reykjavik, Iceland, from 1998 to 2016. Among 1,515 erythromycin-resistant isolates, 90.3% were of only three emm types, emm4 (n = 713), emm6 (n = 324), and emm12 (n = 332), with each being predominant in a distinct epidemic peak. The antibiotic efflux pump genes, mef(A) and msr(D), were present on chimeric mobile genetic elements in 99.3% of the macrolide-resistant isolates of these emm types. Of note, in addition to macrolide resistance, virtually all emm12 isolates had a single amino acid substitution in penicillin-binding protein PBP2X that conferred a 2-fold increased penicillin G and ampicillin MIC among the isolates tested. We conclude that each of the three large epidemic peaks of macrolide-resistant GAS infections occurring in Iceland since 1998 was caused by the emergence and clonal expansion of progenitor strains, with macrolide resistance being conferred predominantly by inducible Mef(A) and Msr(D) drug efflux pumps. The occurrence of emm12 strains with macrolide resistance and decreased beta-lactam susceptibility was unexpected and is of public health concern., This study was supported in part by the Fondren Foundation, Houston Methodist Hospital and Research Institute, and National Institutes of Health grants AI139369 and AI146771 (to J.M.M.).
- Published
- 2020
44. Streptolysin O Rapidly Impairs Neutrophil Oxidative Burst and Antibacterial Responses to Group A Streptococcus.
- Author
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Uchiyama, Satoshi, Döhrmann, Simon, Timmer, Anjuli M., Dixit, Neha, Ghochani, Mariam, Bhandari, Tamara, Timmer, John C., Sprague, Kimberly, Bubeck-Wardenburg, Juliane, Simon, Scott I., Nizet, Victor, Alonzo, Francis, and Beres, Stephen B.
- Subjects
STREPTOCOCCUS ,PHARYNGITIS ,TOXIC shock syndrome - Abstract
Group A Streptococcus (GAS) causes a wide range of human infections, ranging from simple pharyngitis to life-threatening necrotizing fasciitis and toxic shock syndrome. A globally disseminated clone of M1T1 GAS has been associated with an increase in severe, invasive GAS infections in recent decades. The secreted GAS pore-forming toxin streptolysin O (SLO), which induces eukaryotic cell lysis in a cholesterol-dependent manner, is highly upregulated in the GAS M1T1 clone during bloodstream dissemination. SLO is known to promote GAS resistance to phagocytic clearance by neutrophils, a critical first element of host defense against invasive bacterial infection. Here, we examine the role of SLO in modulating specific neutrophil functions during their early interaction with GAS. We find that SLO at subcytotoxic concentrations and early time points is necessary and sufficient to suppress neutrophil oxidative burst, in a manner reversed by free cholesterol and anti-SLO blocking antibodies. In addition, SLO at subcytotoxic concentrations blocked neutrophil degranulation, interleukin-8 secretion and responsiveness, and elaboration of DNA-based neutrophil extracellular traps, cumulatively supporting a key role for SLO in GAS resistance to immediate neutrophil killing. A non-toxic SLO derivate elicits protective immunity against lethal GAS challenge in a murine infection model. We conclude that SLO exerts a novel cytotoxic-independent function at early stages of invasive infections (<30 min), contributing to GAS escape from neutrophil clearance. [ABSTRACT FROM AUTHOR]
- Published
- 2016
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45. Streptolysin O Rapidly Impairs Neutrophil Oxidative Burst and Antibacterial Responses to Group A Streptococcus.
- Author
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Satoshi Uchiyama, Döhrmann, Simon, Timmer, Anjuli M., Dixit, Neha, Ghochani, Mariam, Bhandari, Tamara, Timmer, John C., Sprague, Kimberly, Bubeck-Wardenburg, Juliane, Simon, Scott I., Nizet, Victor, Alonzo, Francis, and Beres, Stephen B.
- Subjects
STREPTOCOCCUS pyogenes ,NEUTROPHILS - Abstract
Group A Streptococcus (GAS) causes a wide range of human infections, ranging from simple pharyngitis to life-threatening necrotizing fasciitis and toxic shock syndrome. A globally disseminated clone of M1T1 GAS has been associated with an increase in severe, invasive GAS infections in recent decades. The secreted GAS pore-forming toxin streptolysin O (SLO), which induces eukaryotic cell lysis in a cholesterol-dependent manner, is highly upregulated in the GAS M1T1 clone during bloodstream dissemination. SLO is known to promote GAS resistance to phagocytic clearance by neutrophils, a critical first element of host defense against invasive bacterial infection. Here, we examine the role of SLO in modulating specific neutrophil functions during their early interaction with GAS. We find that SLO at subcytotoxic concentrations and early time points is necessary and sufficient to suppress neutrophil oxidative burst, in a manner reversed by free cholesterol and anti-SLO blocking antibodies. In addition, SLO at subcytotoxic concentrations blocked neutrophil degranulation, interleukin-8 secretion and responsiveness, and elaboration of DNA-based neutrophil extracellular traps, cumulatively supporting a key role for SLO in GAS resistance to immediate neutrophil killing. A non-toxic SLO derivate elicits protective immunity against lethal GAS challenge in a murine infection model. We conclude that SLO exerts a novel cytotoxic-independent function at early stages of invasive infections (<30 min), contributing to GAS escape from neutrophil clearance. [ABSTRACT FROM AUTHOR]
- Published
- 2015
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46. Postpartum Group A Streptococcus Case Series: Reach Out to Infection Prevention!
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Alexander, Andrew J, Myers, Carol, Beres, Stephen B, Olsen, Randall J, Musser, James M, and Mangino, Julie E
- Abstract
A series of postpartum Streptococcus pyogenes infections prompted an investigation to rule out potential transmission by a health care worker. None of the hospital staff screened were colonized. All isolates were determined to be unrelated by molecular methods, including whole-genome sequencing. Thus, nosocomial transmission was considered unlikely. [ABSTRACT FROM AUTHOR]
- Published
- 2018
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47. Interacting networks of resistance, virulence and core machinery genes identified by genome-wide epistasis analysis
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Stephen D. Bentley, Maiju Pesonen, James M. Musser, Marcin J. Skwark, Paul Turner, Santeri Puranen, Claire Chewapreecha, Julian Parkhill, Nicholas J. Croucher, Stephen B. Beres, Erik Aurell, Yingying Xu, Simon R. Harris, Jukka Corander, Medical Research Council (MRC), Skwark, Marcin J [0000-0002-2022-6766], Croucher, Nicholas J [0000-0001-6303-8768], Puranen, Santeri [0000-0001-6388-7110], Chewapreecha, Claire [0000-0002-1313-4011], Xu, Ying Ying [0000-0002-9096-0552], Turner, Paul [0000-0002-1013-7815], Harris, Simon R [0000-0003-1512-6194], Beres, Stephen B [0000-0003-3041-0185], Parkhill, Julian [0000-0002-7069-5958], Apollo - University of Cambridge Repository, Vanderbilt University, Imperial College London, Department of Computer Science, University of Cambridge, University of Oxford, Wellcome Trust Sanger Institute, Houston Methodist Hospital, Cornell University, Department of Applied Physics, Aalto-yliopisto, Aalto University, Department of Mathematics and Statistics, Jukka Corander / Principal Investigator, and Biostatistics Helsinki
- Subjects
Heredity ,Gene Identification and Analysis ,STREPTOCOCCUS-PNEUMONIAE ,Pathology and Laboratory Medicine ,0302 clinical medicine ,112 Statistics and probability ,1183 Plant biology, microbiology, virology ,Genetics & Heredity ,education.field_of_study ,1184 Genetics, developmental biology, physiology ,PENICILLIN RESISTANCE ,Nucleotide Mapping ,ASSOCIATION ,Genomics ,Aminoacyltransferases ,Bacterial Pathogens ,DIRECT-COUPLING ANALYSIS ,Fitness Epistasis ,Medical Microbiology ,Perspective ,Network Analysis ,Genotype ,Bioinformatics ,Locus (genetics) ,Microbial Genomics ,Microbial Sensitivity Tests ,beta-Lactams ,Microbiology ,beta-Lactam Resistance ,03 medical and health sciences ,Bacterial Proteins ,Genome-Wide Association Studies ,Genetics ,Bacterial Genetics ,Humans ,Penicillin-Binding Proteins ,CONTACT PREDICTION ,education ,Molecular Biology Techniques ,Microbial Pathogens ,Molecular Biology ,Ecology, Evolution, Behavior and Systematics ,ta113 ,0604 Genetics ,Science & Technology ,Bacteria ,Gene Mapping ,Organisms ,Computational Biology ,Biology and Life Sciences ,Epistasis, Genetic ,Split-Decomposition Method ,Yeast ,030104 developmental biology ,Genetics, Population ,Genetic Loci ,Mutation ,Peptidyl Transferases ,Epistasis ,Structural Genomics ,030217 neurology & neurosurgery ,Developmental Biology ,0301 basic medicine ,Cancer Research ,Linkage disequilibrium ,Genetic Networks ,Genome ,Database and Informatics Methods ,Medicine and Health Sciences ,Gene Regulatory Networks ,Genetics (clinical) ,Bacterial Genomics ,Microbial Genetics ,Pneumococcus ,Anti-Bacterial Agents ,Deletion Mutation ,Streptococcus pneumoniae ,Group A streptococci ,Pathogens ,Sequence Analysis ,Life Sciences & Biomedicine ,Research Article ,Computer and Information Sciences ,Multiple Alignment Calculation ,lcsh:QH426-470 ,PROTEINS ,Streptococcus pyogenes ,Population ,Epistasis and functional genomics ,Computational biology ,Biology ,Research and Analysis Methods ,Polymorphism, Single Nucleotide ,Computational Techniques ,Selection, Genetic ,Gene ,COEVOLUTION ,Fungi ,Streptococcus ,Human Genetics ,Bacteriology ,Genome Analysis ,EVOLUTION ,lcsh:Genetics ,RESIDUE CONTACTS ,STATISTICAL-METHODS ,Sequence Alignment ,Genome, Bacterial ,Genome-Wide Association Study - Abstract
Recent advances in the scale and diversity of population genomic datasets for bacteria now provide the potential for genome-wide patterns of co-evolution to be studied at the resolution of individual bases. Here we describe a new statistical method, genomeDCA, which uses recent advances in computational structural biology to identify the polymorphic loci under the strongest co-evolutionary pressures. We apply genomeDCA to two large population data sets representing the major human pathogens Streptococcus pneumoniae (pneumococcus) and Streptococcus pyogenes (group A Streptococcus). For pneumococcus we identified 5,199 putative epistatic interactions between 1,936 sites. Over three-quarters of the links were between sites within the pbp2x, pbp1a and pbp2b genes, the sequences of which are critical in determining non-susceptibility to beta-lactam antibiotics. A network-based analysis found these genes were also coupled to that encoding dihydrofolate reductase, changes to which underlie trimethoprim resistance. Distinct from these antibiotic resistance genes, a large network component of 384 protein coding sequences encompassed many genes critical in basic cellular functions, while another distinct component included genes associated with virulence. The group A Streptococcus (GAS) data set population represents a clonal population with relatively little genetic variation and a high level of linkage disequilibrium across the genome. Despite this, we were able to pinpoint two RNA pseudouridine synthases, which were each strongly linked to a separate set of loci across the chromosome, representing biologically plausible targets of co-selection. The population genomic analysis method applied here identifies statistically significantly co-evolving locus pairs, potentially arising from fitness selection interdependence reflecting underlying protein-protein interactions, or genes whose product activities contribute to the same phenotype. This discovery approach greatly enhances the future potential of epistasis analysis for systems biology, and can complement genome-wide association studies as a means of formulating hypotheses for targeted experimental work., Author summary Epistatic interactions between polymorphisms in DNA are recognized as important drivers of evolution in numerous organisms. Study of epistasis in bacteria has been hampered by the lack of densely sampled population genomic data, suitable statistical models and inference algorithms sufficiently powered for extremely high-dimensional parameter spaces. We introduce the first model-based method for genome-wide epistasis analysis and use two of the largest available bacterial population genome data sets on Streptococcus pneumoniae (the pneumococcus) and Streptococcus pyogenes (group A Streptococcus) to demonstrate its potential for biological discovery. Our approach reveals interacting networks of resistance, virulence and core machinery genes in the pneumococcus, which highlights putative candidates for novel drug targets. We also discover a number of plausible targets of co-selection in S. pyogenes linked to RNA pseudouridine synthases. Our method significantly enhances the future potential of epistasis analysis for systems biology, and can complement genome-wide association studies as a means of formulating hypotheses for targeted experimental work.
- Published
- 2019
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