scholarly journals Analysis of Invasive NontypeableHaemophilus influenzaeIsolates Reveals Selection for the Expression State of Particular Phase-Variable Lipooligosaccharide Biosynthetic Genes

2019 ◽  
Vol 87 (5) ◽  
Author(s):  
Zachary N. Phillips ◽  
Charles Brizuela ◽  
Amy V. Jennison ◽  
Megan Staples ◽  
Keith Grimwood ◽  
...  
Keyword(s):  
Gene Expression ◽  
Phase Variation ◽  
Phase Variable ◽  
Chronic Infections ◽  
Biosynthetic Genes ◽  
Content Type ◽  
Invasive Infections ◽  
Acetyl Group ◽  
Overt Disease ◽  
Expression Status

ABSTRACTNontypeableHaemophilus influenzae(NTHi) is a major human pathogen, responsible for several acute and chronic infections of the respiratory tract. The incidence of invasive infections caused by NTHi is increasing worldwide. NTHi is able to colonize the nasopharynx asymptomatically, and the exact change(s) responsible for transition from benign carriage to overt disease is not understood. We have previously reported that phase variation (the rapid and reversible ON-OFF switching of gene expression) of particular lipooligosaccharide (LOS) glycosyltransferases occurs during transition from colonizing the nasopharynx to invading the middle ear. Variation in the structure of the LOS is dependent on the ON/OFF expression status of each of the glycosyltransferases responsible for LOS biosynthesis. In this study, we surveyed a collection of invasive NTHi isolates for ON/OFF expression status of seven phase-variable LOS glycosyltransferases. We report that the expression state of the LOS biosynthetic genesoafAON andlic2AOFF shows a correlation with invasive NTHi isolates. We hypothesize that these gene expression changes contribute to the invasive potential of NTHi. OafA expression, which is responsible for the addition of anO-acetyl group onto the LOS, has been shown to impart a phenotype of increased serum resistance and may serve as a marker for invasive NTHi.

scholarly journals Global Changes in Mycoplasma gallisepticum Phase-Variable Lipoprotein GenevlhAExpression duringIn VivoInfection of the Natural Chicken Host

2015 ◽  
Vol 84 (1) ◽  
pp. 351-355 ◽  
Author(s):  
K. Pflaum ◽  
E. R. Tulman ◽  
J. Beaudet ◽  
X. Liao ◽  
S. J. Geary
Keyword(s):  
Gene Expression ◽  
Phase Variation ◽  
Mycoplasma Gallisepticum ◽  
Chronic Respiratory Disease ◽  
Etiologic Agent ◽  
Economic Losses ◽  
Global Changes ◽  
Phase Variable ◽  
Content Type ◽  
Host Interaction

Mycoplasma gallisepticumis the primary etiologic agent of chronic respiratory disease in poultry, a disease largely affecting the respiratory tract and causing significant economic losses worldwide. Immunodominant proteins encoded by members of the variable lipoprotein and hemagglutinin (vlhA) gene family are thought to be important for mechanisms ofM. gallisepticum-host interaction, pathogenesis, and immune evasion, but their exact role and the overall nature of their phase variation are unknown. To better understand these mechanisms, we assessed global transcriptomicvlhAgene expression directly fromM. gallisepticumpopulations present on tracheal mucosae during a 7-day experimental infection in the natural chicken host. Here we report differences in both dominant and minorvlhAgene expression levels throughout the first week of infection and starting as early as day 1 postinfection, consistent with a functional role not dependent on adaptive immunity for driving phase variation. Notably, data indicated that, at given time points, specificvlhAgenes were similarly dominant in multiple independent hosts, suggesting a nonstochastic temporal progression of dominantvlhAgene expression in the colonizing bacterial population. The dominant expression of a givenvlhAgene was not dependent on the presence of 12-copy GAA trinucleotide repeats in the promoter region and did not revert to the predominatevlhAgene when no longer faced with host pressures. Overall, these data indicate thatvlhAphase variation is dynamic throughout the earliest stages of infection and that the pattern of dominantvlhAexpression may be nonrandom and regulated by previously unrecognized mechanisms.

scholarly journals Phasevarions Mediate Epigenetic Regulation of Antimicrobial Susceptibility in Neisseria meningitidis

2014 ◽  
Vol 58 (7) ◽  
pp. 4219-4221 ◽  
Author(s):  
Freda E.-C. Jen ◽  
Kate L. Seib ◽  
Michael P. Jennings
Keyword(s):  
Gene Expression ◽  
Neisseria Meningitidis ◽  
Epigenetic Regulation ◽  
Antimicrobial Susceptibility ◽  
Bacterial Pathogens ◽  
Phase Variation ◽  
Dna Methyltransferases ◽  
Global Methylation ◽  
Content Type ◽  
Multiple Genes

ABSTRACTPhase variation is a common feature of host-adapted bacterial pathogens such asNeisseria meningitidis. Recently, we reported that this rapid on/off switching of gene expression occurs in DNA methyltransferases, altering expression in multiple genes via changes in global methylation. In the current study, we compared MIC values of strains with ModA11, ModA12, and ModD1 phasevarions, revealing MIC differences due to ModA11 and ModA12 switching, with a ModA11_OFF strain showing 4-fold reduced susceptibilities to ceftazidime and ciprofloxacin.

scholarly journals PrtR Homeostasis Contributes to Pseudomonas aeruginosa Pathogenesis and Resistance against Ciprofloxacin

2014 ◽  
Vol 82 (4) ◽  
pp. 1638-1647 ◽  
Author(s):  
Ziyu Sun ◽  
Jing Shi ◽  
Chang Liu ◽  
Yongxin Jin ◽  
Kewei Li ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Bacterial Colonization ◽  
Mrna Level ◽  
Sos Response ◽  
Opportunistic Pathogen ◽  
Host Cells ◽  
Chronic Infections ◽  
Mobility Shift ◽  
Content Type

ABSTRACTPseudomonas aeruginosais an opportunistic pathogen that causes acute and chronic infections in humans. Pyocins are bacteriocins produced byP. aeruginosathat are usually released through lysis of the producer strains. Expression of pyocin genes is negatively regulated by PrtR, which gets cleaved under SOS response, leading to upregulation of pyocin synthetic genes. Previously, we demonstrated that PrtR is required for the expression of type III secretion system (T3SS), which is an important virulence component ofP. aeruginosa. In this study, we demonstrate that mutation inprtRresults in reduced bacterial colonization in a mouse acute pneumonia model. Examination of bacterial and host cells in the bronchoalveolar lavage fluids from infected mice revealed that expression of PrtR is induced by reactive oxygen species (ROS) released by neutrophils. We further demonstrate that treatment with hydrogen peroxide or ciprofloxacin, known to induce the SOS response and pyocin production, resulted in an elevated PrtR mRNA level. Overexpression of PrtR by atacpromoter repressed the endogenousprtRpromoter activity, and electrophoretic mobility shift assay revealed that PrtR binds to its own promoter, suggesting an autorepressive mechanism of regulation. A high level of PrtR expressed from a plasmid resulted in increased T3SS gene expression during infection and higher resistance against ciprofloxacin. Overall, our results suggest that the autorepression of PrtR contributes to the maintenance of a relatively stable level of PrtR, which is permissive to T3SS gene expression in the presence of ROS while increasing bacterial tolerance to stresses, such as ciprofloxacin, by limiting pyocin production.

scholarly journals Genome Sequences of Two Pseudomonas aeruginosa Isolates with Defects in Type III Secretion System Gene Expression from a Chronic Ankle Wound Infection

2021 ◽  
Author(s):  
Sardar Karash ◽  
Robert Nordell ◽  
Egon A. Ozer ◽  
Timothy L. Yahr
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Type Iii Secretion ◽  
Biofilm Formation ◽  
Secretion System ◽  
Chronic Infections ◽  
Genome Sequences ◽  
Content Type ◽  
Chronic Colonization ◽  
Host Tolerance

A common feature of microorganisms that cause chronic infections is a stealthy lifestyle that promotes immune avoidance and host tolerance. During chronic colonization of cystic fibrosis (CF) patients, Pseudomonas aeruginosa acquires numerous adaptations that include reduced expression of some factors, such as motility, O antigen, and the T3SS, and increased expression of other traits, such as biofilm formation.

scholarly journals Stabilizing Genetically Unstable Simple Sequence Repeats in the Campylobacter jejuni Genome by Multiplex Genome Editing: a Reliable Approach for Delineating Multiple Phase-Variable Genes

mBio ◽  
2021 ◽  
Author(s):  
Shouji Yamamoto ◽  
Sunao Iyoda ◽  
Makoto Ohnishi
Keyword(s):  
Campylobacter Jejuni ◽  
Simple Sequence Repeat ◽  
Phase Variation ◽  
Developed Countries ◽  
Phase Variable ◽  
Phenotypic Stability ◽  
Content Type ◽  
Multiple Phase ◽  
Simple Sequence ◽  
Variable Genes

Campylobacter jejuni is the leading bacterial cause of foodborne gastroenteritis in developed countries and occasionally progresses to the autoimmune disease Guillain-Barré syndrome. A relatively large number of hypermutable simple sequence repeat (SSR) tracts in the C. jejuni genome markedly decreases its phenotypic stability through reversible changes in the ON or OFF expression states of the genes in which they reside, a phenomenon called phase variation.

scholarly journals Systematic Analysis of REBASE Identifies Numerous Type I Restriction-Modification Systems with Duplicated, Distinct hsdS Specificity Genes That Can Switch System Specificity by Recombination

mSystems ◽  
2020 ◽  
Vol 5 (4) ◽  
Author(s):  
John M. Atack ◽  
Chengying Guo ◽  
Thomas Litfin ◽  
Long Yang ◽  
Patrick J. Blackall ◽  
...  
Keyword(s):  
Vaccine Development ◽  
Phase Variation ◽  
Dna Methyltransferases ◽  
Inverted Repeats ◽  
Global Changes ◽  
Type I ◽  
Phase Variable ◽  
Global Methylation ◽  
Content Type ◽  
Restriction Modification

ABSTRACT N6-Adenine DNA methyltransferases associated with some Type I and Type III restriction-modification (R-M) systems are able to undergo phase variation, randomly switching expression ON or OFF by varying the length of locus-encoded simple sequence repeats (SSRs). This variation of methyltransferase expression results in genome-wide methylation differences and global changes in gene expression. These epigenetic regulatory systems are called phasevarions, phase-variable regulons, and are widespread in bacteria. A distinct switching system has also been described in Type I R-M systems, based on recombination-driven changes in hsdS genes, which dictate the DNA target site. In order to determine the prevalence of recombination-driven phasevarions, we generated a program called RecombinationRepeatSearch to interrogate REBASE and identify the presence and number of inverted repeats of hsdS downstream of Type I R-M loci. We report that 3.9% of Type I R-M systems have duplicated variable hsdS genes containing inverted repeats capable of phase variation. We report the presence of these systems in the major pathogens Enterococcus faecalis and Listeria monocytogenes, which could have important implications for pathogenesis and vaccine development. These data suggest that in addition to SSR-driven phasevarions, many bacteria have independently evolved phase-variable Type I R-M systems via recombination between multiple, variable hsdS genes. IMPORTANCE Many bacterial species contain DNA methyltransferases that have random on/off switching of expression. These systems, called phasevarions (phase-variable regulons), control the expression of multiple genes by global methylation changes. In every previously characterized phasevarion, genes involved in pathobiology, antibiotic resistance, and potential vaccine candidates are randomly varied in their expression, commensurate with methyltransferase switching. Our systematic study to determine the extent of phasevarions controlled by invertible Type I R-M systems will provide valuable information for understanding how bacteria regulate genes and is key to the study of physiology, virulence, and vaccine development; therefore, it is critical to identify and characterize phase-variable methyltransferases controlling phasevarions.

scholarly journals Bypassing Phase Variation of Lipooligosaccharide (LOS): Using Heptose 1 Glycan Mutants To Establish Widespread Efficacy of Gonococcal Anti-LOS Monoclonal Antibody 2C7

2019 ◽  
Vol 88 (2) ◽  
Author(s):  
Srinjoy Chakraborti ◽  
Sunita Gulati ◽  
Bo Zheng ◽  
Frank J. Beurskens ◽  
Janine Schuurman ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Polymorphonuclear Leukocytes ◽  
Phase Variation ◽  
Human Infection ◽  
Phase Variable ◽  
Content Type ◽  
Human Igg1 ◽  
And Function

ABSTRACT The sialylatable lacto-N-neotetraose (LNnT; Gal-GlcNAc-Gal-Glc) moiety from heptose I (HepI) of the lipooligosaccharide (LOS) of Neisseria gonorrhoeae undergoes positive selection during human infection. Lactose (Gal-Glc) from HepII, although phase variable, is commonly expressed in humans; loss of HepII lactose compromises gonococcal fitness in mice. Anti-LOS monoclonal antibody (MAb) 2C7, a promising antigonococcal immunotherapeutic that elicits complement-dependent bactericidal activity and attenuates gonococcal colonization in mice, recognizes an epitope comprised of lactoses expressed simultaneously from HepI and HepII. Glycan extensions beyond lactose on HepI modulate binding and function of MAb 2C7 in vitro. Here, four gonococcal LOS mutants, each with lactose from HepII but fixed (unable to phase-vary) LOS HepI glycans extended beyond the lactose substitution of HepI (lactose alone, Gal-lactose, LNnT, or GalNAc-LNnT), were used to define how HepI glycan extensions affect (i) mouse vaginal colonization and (ii) efficacy in vitro and in vivo of a human IgG1 chimeric derivative of MAb 2C7 (2C7-Ximab) with a complement-enhancing E-to-G Fc mutation at position 430 (2C7-Ximab-E430G). About 10-fold lower 2C7-Ximab-E430G concentrations achieved similar complement-dependent killing of three gonococcal mutants with glycan extensions beyond lactose-substituted HepI (lactose alone, LNnT, or GalNAc-LNnT) as 2C7-Ximab (unmodified Fc). The fourth mutant (Gal-lactose) resisted direct complement-dependent killing but was killed approximately 70% by 2C7-Ximab-E430G in the presence of polymorphonuclear leukocytes and complement. Only mutants with (sialylatable) LNnT from HepI colonized mice for >3 days, reiterating the importance of LNnT sialylation for infection. 2C7-Ximab-E430G significantly attenuated colonization caused by the virulent mutants.

scholarly journals Streptococcus pneumoniae TIGR4 Phase-Locked Opacity Variants Differ in Virulence Phenotypes

mSphere ◽  
2017 ◽  
Vol 2 (6) ◽  
Author(s):  
Melissa B. Oliver ◽  
Ankita Basu Roy ◽  
Ranjit Kumar ◽  
Elliot J. Lefkowitz ◽  
W. Edward Swords
Keyword(s):  
Gene Expression ◽  
Recent Work ◽  
Bacterial Species ◽  
Phase Variation ◽  
Nasopharyngeal Colonization ◽  
Content Type ◽  
Bacterial Surface ◽  
Variable Expression ◽  
Dna Methylase ◽  
Single Allele

ABSTRACT A growing number of bacterial species undergo epigenetic phase variation due to variable expression or specificity of DNA-modifying enzymes. For pneumococci, this phase variation has long been appreciated as being revealed by changes in colony opacity, which are reflected in changes in expression or accessibility of factors on the bacterial surface. Recent work showed that recombination-generated variation in alleles of the HsdS DNA methylase specificity subunit mediated pneumococcal phase variation. We generated phase-locked populations of S. pneumoniae TIGR4 expressing a single nonvariant hsdS allele and observed significant differences in gene expression and virulence. These results highlight the importance of focused pathogenesis studies within specific phase types. Moreover, the generation of single-allele hsdS constructs will greatly facilitate such studies. Streptococcus pneumoniae (pneumococcus) is a leading human pathogen that can cause serious localized and invasive diseases. Pneumococci can undergo a spontaneous and reversible phase variation that is reflected in colony opacity and which allows the population to adapt to different host environments. Generally, transparent variants are adapted for nasopharyngeal colonization, whereas opaque variants are associated with invasive disease. In recent work, colony phase variation was shown to occur by means of recombination events to generate multiple alleles of the hsdS targeting domain of a DNA methylase complex, which mediates epigenetic changes in gene expression. A panel of isogenic strains were created in the well-studied S. pneumoniae TIGR4 background that are “locked” in the transparent (n = 4) or opaque (n = 2) colony phenotype. The strains had significant differences in colony size which were stable over multiple passages in vitro and in vivo. While there were no significant differences in adherence for the phase-locked mutant strains to immortalized epithelial cells, biofilm formation and viability were reduced for the opaque variants in static assays. Nasopharyngeal colonization was stable for all strains, but the mortality rates differed between them. Transcript profiling by transcriptome sequencing (RNA-seq) analyses revealed that the expression levels of certain virulence factors were increased in a phase-specific manner. As epigenetic regulation of phase variation (often referred to as "phasevarion") is emerging as a common theme for mucosal pathogens, these results serve as a model for future studies of host-pathogen interactions. IMPORTANCE A growing number of bacterial species undergo epigenetic phase variation due to variable expression or specificity of DNA-modifying enzymes. For pneumococci, this phase variation has long been appreciated as being revealed by changes in colony opacity, which are reflected in changes in expression or accessibility of factors on the bacterial surface. Recent work showed that recombination-generated variation in alleles of the HsdS DNA methylase specificity subunit mediated pneumococcal phase variation. We generated phase-locked populations of S. pneumoniae TIGR4 expressing a single nonvariant hsdS allele and observed significant differences in gene expression and virulence. These results highlight the importance of focused pathogenesis studies within specific phase types. Moreover, the generation of single-allele hsdS constructs will greatly facilitate such studies.

scholarly journals Phase Variation of NadA in Invasive Neisseria meningitidis Isolates Impacts on Coverage Estimates for 4C-MenB, a MenB Vaccine

2018 ◽  
Vol 56 (9) ◽  
Author(s):  
Luke R. Green ◽  
Jay Lucidarme ◽  
Neelam Dave ◽  
Hannah Chan ◽  
Stephen Clark ◽  
...  
Keyword(s):  
Neisseria Meningitidis ◽  
High Throughput ◽  
Meningococcal Disease ◽  
Regulatory Region ◽  
Phase Variation ◽  
Enzyme Linked Immunosorbent Assay ◽  
Phase Variable ◽  
Content Type ◽  
Protective Antigens ◽  
Low Expression

ABSTRACT A recombinant NadA protein is one of the four major protective antigens of 4C-MenB (Bexsero), a vaccine developed for serogroup B Neisseria meningitidis (MenB). The meningococcal antigen typing system (MATS) is utilized as a high-throughput assay for assessing the invasive MenB strain coverage of 4C-MenB. Where present, the nadA gene is subject to phase-variable changes in transcription due to a 5′TAAA repeat tract located in a regulatory region. The promoter-containing intergenic region (IGR) sequences and 5′TAAA repeat numbers were determined for 906 invasive meningococcal disease isolates possessing the nadA gene. Exclusion of the 5′TAAA repeats reduced the number of IGR alleles from 82 to 23. Repeat numbers were associated with low and high levels of NadA expression by Western blotting and enzyme-linked immunosorbent assay (ELISA). Low-expression repeat numbers were present in 83% of 179 MenB isolates with NadA-2/3 or NadA-1 peptide variants and 68% of 480 MenW ST-11 complex isolates with NadA-2/3 peptide variants. For isolates with vaccine-compatible NadA variants, 93% of MATS-negative isolates were associated with low-expression repeat numbers, whereas 63% of isolates with MATS relative potency (RP) scores above the 95% confidence interval for the positive bactericidal threshold had high-expression repeat numbers. Analysis of 5′TAAA repeat numbers has potential as a rapid, high-throughput method for assessing strain coverage for the NadA component of 4C-MenB. A key application will be assessing coverage in meningococcal disease cases where confirmation is by PCR only and MATS cannot be applied.

scholarly journals Staphylococcus aureus Responds to Physiologically Relevant Temperature Changes by Altering Its Global Transcript and Protein Profile

mSphere ◽  
2021 ◽  
Vol 6 (2) ◽  
Author(s):  
Raeven A. Bastock ◽  
Emily C. Marino ◽  
Richard E. Wiemels ◽  
Donald L. Holzschu ◽  
Rebecca A. Keogh ◽  
...  
Keyword(s):  
Gene Expression ◽  
Staphylococcus Aureus ◽  
Environmental Conditions ◽  
Large Temperature ◽  
Small Scale ◽  
Content Type ◽  
Temperature Changes ◽  
Invasive Infections ◽  
The Impact ◽  
Definite Temperature

ABSTRACT Staphylococcus aureus is an opportunistic pathogen that colonizes the anterior nares of 30 to 50% of the population. Colonization is most often asymptomatic; however, self-inoculation can give rise to potentially fatal infections of the deeper tissues and blood. Like all bacteria, S. aureus can sense and respond to environmental cues and modify gene expression to adapt to specific environmental conditions. The transition of S. aureus from the nares to the deeper tissues and blood is accompanied by changes in environmental conditions, such as nutrient availability, pH, and temperature. In this study, we perform transcriptomics and proteomics on S. aureus cultures growing at three physiologically relevant temperatures, 34°C (nares), 37°C (body), and 40°C (pyrexia), to determine if small scale, biologically meaningful alterations in temperature impact S. aureus gene expression. Results show that small but definite temperature changes elicit a large-scale restructuring of the S. aureus transcriptome and proteome in a manner that, most often, inversely correlates with increasing temperature. We also provide evidence that a large majority of these changes are modulated at the posttranscriptional level, possibly by sRNA regulatory elements. Phenotypic analyses were also performed to demonstrate that these changes have physiological relevance. Finally, we investigate the impact of temperature-dependent alterations in gene expression on S. aureus pathogenesis and demonstrate decreased intracellular invasion of S. aureus grown at 34°C. Collectively, our results demonstrate that small but biologically meaningful alterations in temperature influence S. aureus gene expression, a process that is likely a major contributor to the transition from a commensal to pathogen. IMPORTANCE Enteric bacterial pathogens, like Escherichia coli, are known to experience large temperature differences as they are transmitted through the fecal oral route. This change in temperature has been demonstrated to influence bacterial gene expression and facilitate infection. Staphylococcus aureus is a human-associated pathogen that can live as a commensal on the skin and nares or cause invasive infections of the deeper tissues and blood. Factors influencing S. aureus nasal colonization are not fully understood; however, individuals colonized with S. aureus are at increased risk of invasive infections through self-inoculation. The transition of S. aureus from the nose (colonization) to the body (infection) is accompanied by a modest but definite temperature increase, from 34°C to 37°C. In this study, we investigate whether these host-associated small temperature changes can influence S. aureus gene expression. Results show widespread changes in the bacterial transcriptome and proteome at three physiologically relevant temperatures (34°C, 37°C, and 40°C).

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