scholarly journals The Type IV Pilin of Burkholderia mallei Is Highly Immunogenic but Fails To Protect against Lethal Aerosol Challenge in a Murine Model

2007 ◽  
Vol 75 (6) ◽  
pp. 3027-3032 ◽  
Author(s):  
Paula J. Fernandes ◽  
Qin Guo ◽  
David M. Waag ◽  
Michael S. Donnenberg
Keyword(s):  
High Titer ◽  
Active Immunization ◽  
Burkholderia Mallei ◽  
Subunit Vaccines ◽  
Passive Protection ◽  
Type Iv ◽  
Type Iv Pilin ◽  
A Subunit ◽  
Pilin Protein

ABSTRACT Burkholderia mallei is the cause of glanders and a proven biological weapon. We identified and purified the type IV pilin protein of this organism to study its potential as a subunit vaccine. We found that purified pilin was highly immunogenic. Furthermore, mice infected via sublethal aerosol challenge developed significant increases in titers of antibody against the pilin, suggesting that it is expressed in vivo. Nevertheless, we found no evidence that high-titer antipilin antisera provided passive protection against a sublethal or lethal aerosol challenge and no evidence of protection afforded by active immunization with purified pilin. These results contrast with the utility of type IV pilin subunit vaccines against other infectious diseases and highlight the need for further efforts to identify protective responses against this pathogen.

scholarly journals Biofilms Formed by Nontypeable Haemophilus influenzae In Vivo Contain both Double-Stranded DNA and Type IV Pilin Protein

2007 ◽  
Vol 189 (10) ◽  
pp. 3868-3875 ◽  
Author(s):  
Joseph A. Jurcisek ◽  
Lauren O. Bakaletz
Keyword(s):  
Haemophilus Influenzae ◽  
Outer Membrane Proteins ◽  
Mammalian Host ◽  
Type Iv ◽  
Double Stranded Dna ◽  
Type Iv Pilin ◽  
Pilin Protein ◽  
Biofilm Matrix

ABSTRACT Nontypeable Haemophilus influenzae (NTHI) strains are members of the normal human nasopharyngeal flora, as well as frequent opportunistic pathogens of both the upper and lower respiratory tracts. Recently, it has been shown that NTHI can form biofilms both in vitro and in vivo. NTHI strains within in vitro-formed biofilms differentially express both epitopes of lipooligosaccharide (LOS) and the outer membrane proteins P2, P5, and P6, whereas those generated either in a 96-well plate assay in vitro or in a mammalian host have been shown to incorporate a specific glycoform of sialylated LOS within the biofilm matrix. While DNA has been identified as a key component of the biofilm matrix formed in vitro by several bacterial pathogens, here we demonstrate for the first time that in addition to sialylated LOS, the biofilm formed by NTHI in vivo contains both type IV pilin protein and a significant amount of double-stranded DNA. The DNA appeared to be arranged in a dense interlaced meshwork of fine strands as well as in individual thicker “ropes” that span water channels, suggesting that DNA could be imparting structural stability to the biofilm produced by NTHI in vivo. The presence of type IV pilin protein both appearing as small aggregates within the biofilm matrix and tracking along DNA strands supports our observations which showed that type IV pili are expressed by NTHI during experimental otitis media when these bacteria form a biofilm in the middle ear space.

scholarly journals Nasopharyngeal infection byStreptococcus pyogenesrequires superantigen-responsive Vβ-specific T cells

2017 ◽  
Vol 114 (38) ◽  
pp. 10226-10231 ◽  
Author(s):  
Joseph J. Zeppa ◽  
Katherine J. Kasper ◽  
Ivor Mohorovic ◽  
Delfina M. Mazzuca ◽  
S. M. Mansour Haeryfar ◽  
...  
Keyword(s):  
T Cells ◽  
High Titer ◽  
Passive Immunization ◽  
Cell Receptor ◽  
Active Immunization ◽  
Wild Type ◽  
Toxic Shock ◽  
Variable Domains ◽  
Β Chain

The globally prominent pathogenStreptococcus pyogenessecretes potent immunomodulatory proteins known as superantigens (SAgs), which engage lateral surfaces of major histocompatibility class II molecules and T-cell receptor (TCR) β-chain variable domains (Vβs). These interactions result in the activation of numerous Vβ-specific T cells, which is the defining activity of a SAg. Although streptococcal SAgs are known virulence factors in scarlet fever and toxic shock syndrome, mechanisms by how SAgs contribute to the life cycle ofS. pyogenesremain poorly understood. Herein, we demonstrate that passive immunization against the Vβ8-targeting SAg streptococcal pyrogenic exotoxin A (SpeA), or active immunization with either wild-type or a nonfunctional SpeA mutant, protects mice from nasopharyngeal infection; however, only passive immunization, or vaccination with inactive SpeA, resulted in high-titer SpeA-specific antibodies in vivo. Mice vaccinated with wild-type SpeA rendered Vβ8+T cells poorly responsive, which prevented infection. This phenotype was reproduced with staphylococcal enterotoxin B, a heterologous SAg that also targets Vβ8+T cells, and rendered mice resistant to infection. Furthermore, antibody-mediated depletion of T cells prevented nasopharyngeal infection byS. pyogenes, but not byStreptococcus pneumoniae, a bacterium that does not produce SAgs. Remarkably, these observations suggest thatS. pyogenesuses SAgs to manipulate Vβ-specific T cells to establish nasopharyngeal infection.

scholarly journals Functional Consequences of Sequence Variation in Bundlin, the Enteropathogenic Escherichia coli Type IV Pilin Protein

2007 ◽  
Vol 75 (10) ◽  
pp. 4687-4696 ◽  
Author(s):  
Paula J. Fernandes ◽  
Qin Guo ◽  
Michael S. Donnenberg
Keyword(s):  
Escherichia Coli ◽  
Immune Responses ◽  
Sequence Divergence ◽  
Cross Reactivity ◽  
Human Antibody ◽  
Type Iv ◽  
Volunteer Study ◽  
Pilus Biogenesis ◽  
Type Iv Pilin ◽  
Pilin Protein

ABSTRACT The bundle-forming pilus (BFP) of enteropathogenic Escherichia coli (EPEC) is an important virulence factor. We examined the role of divergent alleles of bfpA encoding bundlin, the BFP pilin protein, in pilus biogenesis, pilus interactions, and immune responses. We found that the BFP biogenesis machine from an EPEC strain that expresses one bundlin type is capable of assembling all other bundlin types. Furthermore, we found that EPEC strains expressing divergent bundlin types are capable of forming mixed autoaggregates, suggesting that different pilin types can intertwine. However, we found that there was a marked difference between alleles in immunogenicity in both rabbits and mice of a peptide derived from a region of bundlin undergoing apparent diversifying selection. In addition, despite a high degree of cross-reactivity between divergent bundlin proteins, in both mice and rabbits responses appeared to be stronger against the homologous pilin protein than against the heterologous protein. This result was verified using sera from a volunteer study, which demonstrated that the human antibody responses after an initial challenge with live EPEC were stronger against the homologous bundlin protein than against a divergent bundlin protein. However, a repeat challenge induced equivalent responses. These results are consistent with the hypothesis that human immune responses against bundlin exert selective pressure on bfpA sequence divergence.

scholarly journals A Type IV Pilin, PilA, Contributes to Adherence of Burkholderia pseudomallei and Virulence In Vivo

2005 ◽  
Vol 73 (2) ◽  
pp. 1260-1264 ◽  
Author(s):  
Angela E. Essex-Lopresti ◽  
Justin A. Boddey ◽  
Richard Thomas ◽  
Martin P. Smith ◽  
M. Gill Hartley ◽  
...  
Keyword(s):  
Murine Model ◽  
Burkholderia Pseudomallei ◽  
Deletion Mutant ◽  
Type Iv Pili ◽  
Structural Protein ◽  
Type Iv ◽  
Type Iv Pilin ◽  
Reduced Adherence ◽  
Multiple Type

ABSTRACT The Burkholderia pseudomallei K96243 genome contains multiple type IV pilin-associated loci, including one encoding a putative pilus structural protein (pilA). A pilA deletion mutant has reduced adherence to human epithelial cells and is less virulent in the nematode model of virulence and the murine model of melioidosis, suggesting a role for type IV pili in B. pseudomallei virulence.

scholarly journals Expression of a Clostridium perfringens Type IV Pilin by Neisseria gonorrhoeae Mediates Adherence to Muscle Cells

2011 ◽  
Vol 79 (8) ◽  
pp. 3096-3105 ◽  
Author(s):  
Katherine Rodgers ◽  
Cindy Grove Arvidson ◽  
Stephen Melville
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Clostridium Perfringens ◽  
Klebsiella Oxytoca ◽  
Wild Type ◽  
Unique Type ◽  
Gram Negative ◽  
Content Type ◽  
Type Iv ◽  
Type Iv Pilin

ABSTRACTClostridium perfringensis an anaerobic, Gram-positive bacterium that causes a range of diseases in humans, including lethal gas gangrene. We have recently shown that strains ofC. perfringensmove across the surface of agar plates by a unique type IV pilus (TFP)-mediated social motility that had not been previously described. Based on sequence homology to pilins in Gram-negative bacteria,C. perfringensappears to have two pilin subunits, PilA1 and PilA2. Structural prediction analysis indicated PilA1 is similar to the pseudopilin found inKlebsiella oxytoca, while PilA2 is more similar to true pilins found in the Gram-negative pathogensPseudomonas aeruginosaandNeisseria gonorrhoeae. Strains ofN. gonorrhoeaethat were genetically deficient in the native pilin, PilE, but supplemented with inducible expression of PilA1 and PilA2 ofC. perfringenswere constructed. Genetic competence, wild-type twitching motility, and attachment to human urogenital epithelial cells were not restored by expression of either pilin. However, attachment to mouse and rat myoblast (muscle) cell lines was observed with theN. gonorrhoeaestrain expressing PilA2. Significantly, wild-typeC. perfringenscells adhered to mouse myoblasts under anaerobic conditions, and adherence was 10-fold lower in apilTmutant that lacked functional TFP. These findings implicateC. perfringensTFP in the ability ofC. perfringensto adhere to and move along muscle fibersin vivo, which may provide a therapeutic approach to limiting this rapidly spreading and highly lethal infection.

scholarly journals Production of pilus-like filaments in Geobacter sulfurreducens in the absence of the type IV pilin protein PilA

2010 ◽  
Vol 310 (1) ◽  
pp. 62-68 ◽  
Author(s):  
Anna Klimes ◽  
Ashley E. Franks ◽  
Richard H. Glaven ◽  
Hoa Tran ◽  
Christian L. Barrett ◽  
...  
Keyword(s):  
Geobacter Sulfurreducens ◽  
Type Iv ◽  
Type Iv Pilin ◽  
Pilin Protein

scholarly journals Molecular Variation among Type IV Pilin (bfpA) Genes from Diverse Enteropathogenic Escherichia coli Strains

2000 ◽  
Vol 68 (12) ◽  
pp. 7028-7038 ◽  
Author(s):  
T. Eric Blank ◽  
Hailang Zhong ◽  
Alison L. Bell ◽  
Thomas S. Whittam ◽  
Michael S. Donnenberg
Keyword(s):  
Escherichia Coli ◽  
Host Immune System ◽  
Enzyme Electrophoresis ◽  
Sequence Comparisons ◽  
Variable Regions ◽  
Type Iv ◽  
Pcr Products ◽  
Type Iv Pilin ◽  
Single Base Pair ◽  
Pilin Protein

ABSTRACT Typical enteropathogenic Escherichia coli (EPEC) strains produce bundle-forming pili (BFP), type IVB fimbriae that have been implicated in EPEC virulence, antigenicity, autoaggregation, and localized adherence to epithelial cells (LA). BFP are polymers of bundlin, a pilin protein that is encoded by the bfpA gene found on a large EPEC plasmid. Striking sequence variation has previously been observed among type IV pilin genes of other gram-negative bacterial pathogens (e.g., Pseudomonas andNeisseria spp.). In contrast, the established sequences ofbfpA genes from two distantly related prototype EPEC strains vary by only a single base pair. To determine whether bundlin sequences vary more extensively, we used PCR to amplify thebfpA genes from 19 EPEC strains chosen for their various serotypes and sites and years of isolation. Eight differentbfpA alleles were identified by sequencing of the PCR products. These alleles can be classified into two major groups. The α group contains three alleles derived from strains carrying O55, O86, O111, O119, O127, or O128 somatic antigens. The β group contains five alleles derived from strains carrying O55, O110, O128ab, O142, or nontypeable antigens. Sequence comparisons show that bundlin has highly conserved and variable regions, with most of the variation occurring in the C-terminal two-thirds of the protein. The results of multilocus enzyme electrophoresis support the hypothesis that bfpAsequences have spread horizontally across distantly related clonal lineages. Strains with divergent bundlin sequences express bundlin protein, produce BFP, and carry out autoaggregation and LA. However, four strains lack most or all of these phenotypes despite having an intact bfpA gene. These results have important implications for our understanding of bundlin structure, transmission of thebfp gene cluster among EPEC strains, and the role of bundlin variation in the evasion of host immune system responses.

Absence of high molecular weight proteins 1 and/or 2 is associated with decreased adherence among non-typeable Haemophilus influenzae clinical isolates

2013 ◽  
Vol 62 (11) ◽  
pp. 1649-1656 ◽  
Author(s):  
Jeni Vuong ◽  
Xin Wang ◽  
Jordan M. Theodore ◽  
Jennifer Whitmon ◽  
Patricia Gomez de Leon ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Haemophilus Influenzae ◽  
High Molecular Weight ◽  
Surface Proteins ◽  
Clinical Isolates ◽  
Type Iv ◽  
High Adherence ◽  
Genes Encoding ◽  
Type Iv Pilin ◽  
Pilin Protein

High molecular weight (Hmw) proteins 1 and 2, type IV pilin protein (PilA), outer-membrane protein P5 (OmpP5), Haemophilus protein D (Hpd) and Haemophilus adhesive protein (Hap) are surface proteins involved in the adherence of non-typeable Haemophilus influenzae. One hundred clinical isolates were evaluated for the presence of the genes encoding these proteins by PCR and for their adherence capacity (AC) to Detroit 562 nasopharyngeal cells (D562). The majority of isolates were from blood (77/100); other sites were also represented. Confluent D562 monolayers (1.2×105 cells per well) were inoculated with standardized minimal infective doses (m.o.i.) of 102, 103 or 104 c.f.u. per well. The AC was categorized as low (<10 %) or high (≥10 %) depending on the percentage of c.f.u. adhering per well. All the isolates evaluated showed adherence: 69/100 (69 %) demonstrated high adherence, while 31/100 (31 %) showed low adherence. Of all the genes evaluated, hmw1A and/or hmw2A were detected in 69/100 (69 %) of isolates. The presence of hmw1A and/or hmw2A was associated with increased adherence to D562 cells (P≤0.001). Dot immunoblots were performed to detect protein expression using mAbs 3D6, AD6 and 10C5. Among the high-adherence isolates (n = 69), 72 % reacted with 3D6 and 21 % with 10C5. Our data indicate that the absence of Hmw1 and/or Hmw2 was associated with decreased adherence to D562 cells.

scholarly journals Dual-Function Vaccine for Pseudomonas aeruginosa: Characterization of Chimeric Exotoxin A-Pilin Protein

2001 ◽  
Vol 69 (11) ◽  
pp. 6962-6969 ◽  
Author(s):  
Ralf Hertle ◽  
Randall Mrsny ◽  
David J. Fitzgerald
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Chimeric Protein ◽  
Infectious Agent ◽  
Bacterial Adherence ◽  
Dual Function ◽  
Exotoxin A ◽  
Type Iv ◽  
Type Iv Pilin ◽  
Immunological Assays ◽  
Pilin Protein

ABSTRACT Pseudomonas aeruginosa is the major infectious agent of concern for cystic fibrosis patients. Strategies to prevent colonization by this bacterium and/or neutralize its virulence factors are clearly needed. Here we characterize a dual-function vaccine designed to generate antibodies to reduce bacterial adherence and to neutralize the cytotoxic activity of exotoxin A. To construct the vaccine, key sequences from type IV pilin were inserted into a vector encoding a nontoxic (active-site deletion) version of exotoxin A. The chimeric protein, termed PE64Δ553pil, was expressed inEscherichia coli, refolded to a near-native conformation, and then characterized by various biochemical and immunological assays. PE64Δ553pil bound specifically to asialo-GM1, and, when injected into rabbits, produced antibodies that reduced bacterial adherence and neutralized the cell-killing activity of exotoxin A. Results support further evaluation of this chimeric protein as a vaccine to prevent Pseudomonascolonization in susceptible individuals.

scholarly journals Natural Competence for DNA Transformation by Legionella pneumophila and Its Association with Expression of Type IV Pili

1999 ◽  
Vol 181 (5) ◽  
pp. 1395-1402 ◽  
Author(s):  
Barbara J. Stone ◽  
Yousef Abu Kwaik
Keyword(s):  
Legionella Pneumophila ◽  
Plasmid Dna ◽  
Mammalian Cells ◽  
Type Iv Pili ◽  
Dna Transformation ◽  
Dna Uptake ◽  
Chromosomal Dna ◽  
Type Iv ◽  
Type Iv Pilin ◽  
Pilin Protein

ABSTRACT We have recently described the expression of two pili of different lengths on the surface of Legionella pneumophila (B. J. Stone and Y. Abu Kwaik, Infect. Immun. 66:1768–1775, 1998). Production of long pili requires a functional pilE L locus, encoding a type IV pilin protein. Since type IV pili in Neisseria gonorrhoeaeare associated with competence for DNA transformation, we examined the competence of L. pneumophila for DNA transformation under conditions that allowed the expression of type IV pili. We show that L. pneumophila is naturally competent for DNA transformation by isogenic chromosomal DNA and by plasmid DNA containing L. pneumophila DNA. Many different L. pneumophila loci are able to transform L. pneumophilaafter addition of plasmid DNA, including gspA,ppa, asd, and pilE L. The transformation frequency is reduced when competing DNA containing either L. pneumophila DNA or vector sequences is added to the bacteria, suggesting that uptake-specific sequences may not be involved in DNA uptake. Competence for DNA transformation correlates with expression of the type IV pili, and apilE L mutant defective in expression of type IV pili is not competent for DNA transformation. Complementation of the mutant for competence is restored by the reintroduction of a cosmid that restores production of type IV pili. Minimal competence is restored to the mutant by introduction of pilE Lalone. We conclude that competence for DNA transformation in L. pneumophila is associated with expression of the type IV pilus and results in recombination of L. pneumophila DNA into the chromosome. Since expression of type IV pili also facilitates attachment of L. pneumophila to mammalian cells and protozoa, we designated the type IV pili CAP (for competence- and adherence-associated pili).

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