scholarly journals Expression Microarray and Mouse Virulence Analysis of Four Conserved Two-Component Gene Regulatory Systems in Group A Streptococcus

2006 ◽  
Vol 74 (2) ◽  
pp. 1339-1351 ◽  
Author(s):  
Izabela Sitkiewicz ◽  
James M. Musser
Keyword(s):  
Mutant Strain ◽  
Group A Streptococcus ◽  
Expression Microarray ◽  
Regulatory Systems ◽  
Significant Difference ◽  
Two Component ◽  
Mutant Strains ◽  
Group A ◽  
Gene Regulatory ◽  
Component Gene

ABSTRACT Group A streptococcus (GAS) is a gram-positive human bacterial pathogen that causes diseases ranging from relatively mild epithelial cell surface infections to life-threatening invasive episodes. Much is known about the extracellular molecules that contribute to host-pathogen interactions, but in contrast, far less information is available about regulatory genes that control the expression of individual or multiple GAS virulence factors. The eight GAS genomes that have been sequenced have 12 conserved two-component gene regulatory systems (TCSs), but only 3 of these 12 have been studied in detail. Using an allelic replacement strategy with a nonpolar cassette, we inactivated the response regulator of four TCSs that have only weak homology with TCS genes of known or inferred function in other bacteria. The mutant strains were analyzed by expression microarray analysis at four time points and tested in two mouse infection models. Each TCS influenced expression (directly or indirectly) of 12 to 41% of all chromosomal genes, as assessed by growth in Todd-Hewitt broth and a custom Affymetrix GeneChip. None of the isogenic mutant strains was significantly altered for mouse virulence based on intraperitoneal inoculation. Similarly, compared to the wild-type strain, there was no significant difference in skin lesion size for three of the four mutants. In contrast, the ΔM5005_Spy_0680 mutant strain produced significantly larger abscesses after subcutaneous inoculation into mice, consistent with a hypervirulence phenotype. The mutant strain had significantly higher in vitro expression of several proven and putative virulence genes, including scpA, encoding a peptidase that inactivates complement protein C5a. Together, the data provide new information about previously uncharacterized GAS TCSs.

scholarly journals Yersinia pestis Two-Component Gene Regulatory Systems Promote Survival in Human Neutrophils

2009 ◽  
Vol 78 (2) ◽  
pp. 773-782 ◽  
Author(s):  
Jason L. O'Loughlin ◽  
Justin L. Spinner ◽  
Scott A. Minnich ◽  
Scott D. Kobayashi
Keyword(s):  
Immune System ◽  
Yersinia Pestis ◽  
Innate Immune System ◽  
Intracellular Survival ◽  
Innate Immune ◽  
Human Neutrophils ◽  
Regulatory Systems ◽  
Two Component ◽  
Gene Regulatory ◽  
Component Gene

ABSTRACT Human polymorphonuclear leukocytes (PMNs, or neutrophils) are the most abundant innate immune cell and kill most invading bacteria through combined activities of reactive oxygen species (ROS) and antimicrobial granule constituents. Pathogens such as Yersinia pestis resist destruction by the innate immune system and are able to survive in macrophages and neutrophils. The specific molecular mechanisms used by Y. pestis to survive following phagocytosis by human PMNs are incompletely defined. To gain insight into factors that govern Y. pestis intracellular survival in neutrophils, we inactivated 25 two-component gene regulatory systems (TCSs) with known or inferred function and assessed susceptibility of these mutant strains to human PMN granule extracts. Y. pestis strains deficient for PhoPQ, KdpED, CheY, CvgSY, and CpxRA TCSs were selected for further analysis, and all five strains were altered for survival following interaction with PMNs. Of these five strains, only Y. pestis ΔphoPQ demonstrated global sensitivity to a panel of seven individual neutrophil antimicrobial peptides and serine proteases. Notably, Y. pestis ΔphoPQ was deficient for intracellular survival in PMNs. Iterative analysis with Y. pestis strains lacking the PhoP-regulated genes ugd and pmrK indicated that the mechanism most likely responsible for increased resistance to killing is 4-amino-4-deoxy-l-arabinose modification of lipid A. Together, the data provide new information about Y. pestis evasion of the innate immune system.

scholarly journals A Two-Component Regulatory System Impacts Extracellular Membrane-Derived Vesicle Production in Group A Streptococcus

mBio ◽  
2016 ◽  
Vol 7 (6) ◽  
Author(s):  
Ulrike Resch ◽  
James Anthony Tsatsaronis ◽  
Anaïs Le Rhun ◽  
Gerald Stübiger ◽  
Manfred Rohde ◽  
...  
Keyword(s):  
Lipid Composition ◽  
Secretory Pathway ◽  
Group A Streptococcus ◽  
Regulatory System ◽  
Surface Proteins ◽  
Streptococcal Toxic Shock Syndrome ◽  
Gram Positive ◽  
Gram Positive Bacteria ◽  
Two Component ◽  
Group A

ABSTRACT Export of macromolecules via extracellular membrane-derived vesicles (MVs) plays an important role in the biology of Gram-negative bacteria. Gram-positive bacteria have also recently been reported to produce MVs; however, the composition and mechanisms governing vesiculogenesis in Gram-positive bacteria remain undefined. Here, we describe MV production in the Gram-positive human pathogen group A streptococcus (GAS), the etiological agent of necrotizing fasciitis and streptococcal toxic shock syndrome. M1 serotype GAS isolates in culture exhibit MV structures both on the cell wall surface and in the near vicinity of bacterial cells. A comprehensive analysis of MV proteins identified both virulence-associated protein substrates of the general secretory pathway in addition to “anchorless surface proteins.” Characteristic differences in the contents, distributions, and fatty acid compositions of specific lipids between MVs and GAS cell membrane were also observed. Furthermore, deep RNA sequencing of vesicular RNAs revealed that GAS MVs contained differentially abundant RNA species relative to bacterial cellular RNA. MV production by GAS strains varied in a manner dependent on an intact two-component system, CovRS, with MV production negatively regulated by the system. Modulation of MV production through CovRS was found to be independent of both GAS cysteine protease SpeB and capsule biosynthesis. Our data provide an explanation for GAS secretion of macromolecules, including RNAs, lipids, and proteins, and illustrate a regulatory mechanism coordinating this secretory response. IMPORTANCE Group A streptococcus (GAS) is a Gram-positive bacterial pathogen responsible for more than 500,000 deaths annually. Establishment of GAS infection is dependent on a suite of proteins exported via the general secretory pathway. Here, we show that GAS naturally produces extracellular vesicles with a unique lipid composition that are laden with proteins and RNAs. Interestingly, both virulence-associated proteins and RNA species were found to be differentially abundant in vesicles relative to the bacteria. Furthermore, we show that genetic disruption of the virulence-associated two-component regulator CovRS leads to an increase in vesicle production. This study comprehensively describes the protein, RNA, and lipid composition of GAS-secreted MVs and alludes to a regulatory system impacting this process.

scholarly journals 453. High Burden of Invasive and Severe Group A Streptococcus Disease Among Native Americans on the White Mountain Apache Tribal Lands

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S223-S223
Author(s):  
Catherine Sutcliffe ◽  
Ryan M Close ◽  
Anne M Davidson ◽  
Angelina Reid ◽  
Dianna Quay ◽  
...  
Keyword(s):  
Native American ◽  
Native Americans ◽  
Group A Streptococcus ◽  
Penetrating Trauma ◽  
The United States ◽  
Incidence Rates ◽  
Significant Difference ◽  
White Mountain Apache ◽  
Group A ◽  
Tribal Lands

Abstract Background Native Americans are overrepresented in outbreaks of Group A Streptococcus (GAS) in the United States (US). In 2016, several invasive cases of GAS were detected at the Whiteriver Indian Health Service (IHS) Hospital in Arizona that primarily serves the White Mountain Apache (WMA) Tribe. The objective of this study was to determine the burden of invasive and severe GAS disease among Native Americans on the WMA Tribal lands. Methods Prospective population and laboratory-based surveillance for invasive and severe GASinfections was conducted for two years from March 2017 through February 2019. A case was defined as a Native American individual living on or around WMA Tribal lands with GAS isolated from a normally sterile body site (invasive) or from a non-sterile site (e.g., wound, throat, ear) requiring hospitalization (severe). Incidence rates were calculated using the IHS User Population as the denominators. Age-standardized incidence rates were calculated using US Census data from 2015 as the reference group. Results 157 cases were identified (Year 1: 85; Year 2: 72), including 42 (27%) invasive and 115 (73%) severe cases. Most cases were adults (88.5%; median age: 40.5 years) and had ≥1 underlying medical condition (99.4%), including alcoholism (57.1%), hypertension (37.2%), and diabetes (34.0%). 47.8% of cases had a trigger in the past two weeks, including penetrating trauma (31.8%) and blunt force trauma (14.0%). For 72.9% of cases, a co-infection was detected (most commonly Staphylocccus aureus: 96.8%). 4.5% of cases required amputation and 1.9% died within 30 days of initial culture. The incidence of invasive and severe GAS was 460.9 per 100,000 persons (95% confidence interval: 394.3, 538.8), with no significant difference by year. The incidence was highest among adults ≥65 and lowest among children 5–17 years of age. Age-standardized incidence rates of invasive and severe GAS and invasive only GAS are presented in the Figure. Conclusion The WMA community has experienced disproportionately high rates of invasive and severe GAS for over two years. Studies to determine the reservoirs for transmission are urgently needed, as are interventions to reduce the morbidity and mortality associated with these infections. Disclosures All authors: No reported disclosures.

scholarly journals Group A Streptococcus AdcR Regulon Participates in Bacterial Defense against Host-Mediated Zinc Sequestration and Contributes to Virulence

2020 ◽  
Vol 88 (8) ◽  
Author(s):  
Nishanth Makthal ◽  
Hackwon Do ◽  
Brian M. Wendel ◽  
Randall J. Olsen ◽  
John D. Helmann ◽  
...  
Keyword(s):  
Pathogenic Bacteria ◽  
Group A Streptococcus ◽  
Zn Deficiency ◽  
Direct Competition ◽  
Mutant Strains ◽  
Genes Encoding ◽  
Group A ◽  
Gas Interactions

ABSTRACT Colonization by pathogenic bacteria depends on their ability to overcome host nutritional defenses and acquire nutrients. The human pathogen group A streptococcus (GAS) encounters the host defense factor calprotectin (CP) during infection. CP inhibits GAS growth in vitro by imposing zinc (Zn) limitation. However, GAS counterstrategies to combat CP-mediated Zn limitation and the in vivo relevance of CP-GAS interactions to bacterial pathogenesis remain unknown. Here, we report that GAS upregulates the AdcR regulon in response to CP-mediated Zn limitation. The AdcR regulon includes genes encoding Zn import (adcABC), Zn sparing (rpsN.2), and Zn scavenging systems (adcAII, phtD, and phtY). Each gene in the AdcR regulon contributes to GAS Zn acquisition and CP resistance. The ΔadcC and ΔrpsN.2 mutant strains were the most susceptible to CP, whereas the ΔadcA, ΔadcAII, and ΔphtD mutant strains displayed less CP sensitivity during growth in vitro. However, the ΔphtY mutant strain did not display an increased CP sensitivity. The varied sensitivity of the mutant strains to CP-mediated Zn limitation suggests distinct roles for individual AdcR regulon genes in GAS Zn acquisition. GAS upregulates the AdcR regulon during necrotizing fasciitis infection in WT mice but not in S100a9−/− mice lacking CP. This suggests that CP induces Zn deficiency in the host. Finally, consistent with the in vitro results, several of the AdcR regulon genes are critical for GAS virulence in WT mice, whereas they are dispensable for virulence in S100a9−/− mice, indicating the direct competition for Zn between CP and proteins encoded by the GAS AdcR regulon during infection.

scholarly journals Regulation of Inhibition of Neutrophil Infiltration by the Two-Component Regulatory System CovRS in Subcutaneous Murine Infection with Group A Streptococcus

2013 ◽  
Vol 81 (3) ◽  
pp. 974-983 ◽  
Author(s):  
Jinquan Li ◽  
Hui Zhu ◽  
Wenchao Feng ◽  
Mengyao Liu ◽  
Yingli Song ◽  
...  
Keyword(s):  
Immune Evasion ◽  
Group A Streptococcus ◽  
Synergistic Effects ◽  
Regulatory System ◽  
Neutrophil Infiltration ◽  
Neutrophil Recruitment ◽  
Innate Immune ◽  
Two Component ◽  
Group A ◽  
Innate Immune Evasion

ABSTRACTHypervirulent invasive group A streptococcus (GAS) isolates inhibit neutrophil infiltration more than pharyngitis isolates do, and the molecular basis of this difference is not well understood. This study was designed to first determine whether natural null mutation of the two-component regulatory system CovRS is responsible for the enhancement of the inhibition of neutrophil recruitment seen in hypervirulent GAS. Next, we examined the role of CovRS-regulated interleukin-8/CXC chemokine peptidase (SpyCEP), C5a peptidase (ScpA), and platelet-activating factor acetylhydrolase (SsE) in the enhanced innate immune evasion. Invasive isolate MGAS5005 induces less neutrophil infiltration and produced a greater lesion area than pharyngitis isolate MGAS2221 in subcutaneous infections of mice. It is known that MGAS5005, but not MGAS2221, has a natural 1-bp deletion in thecovSgene. Replacement ofcovSΔ1bpin MGAS5005 with wild-typecovSresulted in the MGAS2221 phenotype. Deletion ofcovSfrom MGAS2221 resulted in the MGAS5005 phenotype. Tests of single, double, and triple deletion mutants of the MGAS5005sse,spyCEP, andscpAgenes found that SsE plays a more important role than SpyCEP and ScpA in the inhibition of neutrophil recruitment and that SsE, SpyCEP, and ScpA do not have synergistic effects on innate immune evasion by MGAS5005. Deletion ofsse, but notspyCEPorscpA, of MGAS2221 enhances neutrophil recruitment. Thus,covSnull mutations can cause substantial inhibition of neutrophil recruitment by enhancing the expression of the chemoattractant-degrading virulence factors, and SsE, but not SpyCEP or ScpA, is required for CovRS-regulated GAS inhibition of neutrophil infiltration.

scholarly journals RivR Is a Negative Regulator of Virulence Factor Expression in Group A Streptococcus

2012 ◽  
Vol 81 (1) ◽  
pp. 364-372 ◽  
Author(s):  
Jeanette Treviño ◽  
Zhuyun Liu ◽  
Tram N. Cao ◽  
Esmeralda Ramirez-Peña ◽  
Paul Sumby
Keyword(s):  
Mutant Strain ◽  
Virulence Factor ◽  
Regulatory Networks ◽  
Group A Streptococcus ◽  
Human Keratinocytes ◽  
Negative Regulator ◽  
Upstream Region ◽  
Content Type ◽  
Factor Expression ◽  
Group A

The bacterial pathogen group AStreptococcus(GAS) causes human diseases ranging from self-limiting pharyngitis (also known as strep throat) to severely invasive necrotizing fasciitis (also known as the flesh-eating syndrome). To control virulence factor expression, GAS utilizes both protein- and RNA-based mechanisms of regulation. Here we report that the transcription factor RivR (RofA-like protein IV) negatively regulates the abundance of mRNAs encoding the hyaluronic acid capsule biosynthesis proteins (hasABC; ∼7-fold) and the protein G-related α2-macroglobulin-binding protein (grab; ∼29-fold). Our data differ significantly from those of a previous study of the RivR regulon. Given thatgrabandhasABCare also negatively regulated by the two-component system CovR/S (controlofvirulence), we tested whether RivR functions through CovR/S. A comparison ofrivandcovsingle and double mutant strains showed that RivR requires CovR activity forgrabandhasABCregulation. Analysis of the upstream region ofrivRidentified a novel promoter the deletion of which reducedrivRmRNA abundance by 70%. ArivRmutant strain had a reduced ability to adhere to human keratinocytes relative to that of the parental and complemented strains, a phenotype that was abolished upon GAS pretreatment with hyaluronidase, highlighting the importance of capsule regulation by RivR during colonization. TherivRmutant strain was also attenuated for virulence in a murine model of bacteremia infection. Thus, we identify RivR as an important regulator of GAS virulence and provide new insight into the regulatory networks controlling virulence factor production in this pathogen.

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