scholarly journals Bacteriophage Sf6 Tailspike Protein for Detection of Shigella flexneri Pathogens

Viruses ◽  
2018 ◽  
Vol 10 (8) ◽  
pp. 431 ◽  
Author(s):  
Sonja Kunstmann ◽  
Tom Scheidt ◽  
Saskia Buchwald ◽  
Alexandra Helm ◽  
Laurence Mulard ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Detection System ◽  
Shigella Flexneri ◽  
Microtiter Plate ◽  
Diagnostic Tools ◽  
Intensity Increase ◽  
O Antigen ◽  
Visible Light Range ◽  
O Antigens ◽  
Tailspike Protein

Bacteriophage research is gaining more importance due to increasing antibiotic resistance. However, for treatment with bacteriophages, diagnostics have to be improved. Bacteriophages carry adhesion proteins, which bind to the bacterial cell surface, for example tailspike proteins (TSP) for specific recognition of bacterial O-antigen polysaccharide. TSP are highly stable proteins and thus might be suitable components for the integration into diagnostic tools. We used the TSP of bacteriophage Sf6 to establish two applications for detecting Shigella flexneri (S. flexneri), a highly contagious pathogen causing dysentery. We found that Sf6TSP not only bound O-antigen of S. flexneri serotype Y, but also the glucosylated O-antigen of serotype 2a. Moreover, mass spectrometry glycan analyses showed that Sf6TSP tolerated various O-acetyl modifications on these O-antigens. We established a microtiter plate-based ELISA like tailspike adsorption assay (ELITA) using a Strep-tag®II modified Sf6TSP. As sensitive screening alternative we produced a fluorescently labeled Sf6TSP via coupling to an environment sensitive dye. Binding of this probe to the S. flexneri O-antigen Y elicited a fluorescence intensity increase of 80% with an emission maximum in the visible light range. The Sf6TSP probes thus offer a promising route to a highly specific and sensitive bacteriophage TSP-based Shigella detection system.

scholarly journals Molecular Modeling of the Shigella flexneri Serogroup 3 and 5 O-Antigens and Conformational Relationships for a Vaccine Containing Serotypes 2a and 3a

Vaccines ◽  
2020 ◽  
Vol 8 (4) ◽  
pp. 643
Author(s):  
Jason Hlozek ◽  
Sara Owen ◽  
Neil Ravenscroft ◽  
Michelle M. Kuttel
Keyword(s):  
Diarrheal Disease ◽  
Shigella Flexneri ◽  
Quadrivalent Vaccine ◽  
Slight Effect ◽  
Work Support ◽  
O Antigen ◽  
The Common ◽  
O Antigens ◽  
Chain Conformations ◽  
Shared Epitopes

The pathogenic bacterium Shigella flexneri is a leading global cause of diarrheal disease. The O-antigen is the primary vaccine target and distinguishes the 30 serotypes reported. Except for serotype 6, all S. flexneri serotypes have a common backbone repeating unit (serotype Y), with variations in substitution creating the various serotypes. A quadrivalent vaccine containing serotypes 2a and 3a (as well as 6 and Shigella sonnei) is proposed to provide broad protection against non-vaccine S. flexneri serotypes through shared epitopes and conformations. Here we model the O-antigen (O-Ag) conformations of serogroups 3 and 5: a continuation of our ongoing systematic study of the S. flexneri O-antigens that began with serogroup 2. Our simulations show that S. flexneri serogroups 2, 3, and 5 all have flexible O-Ags, with substitutions of the backbone altering the chain conformations in different ways. Our analysis suggests three general heuristics for the effects of substitution on the Shigella O-Ag conformations: (1) substitution on rhamnose C reduces the extension of the O-Ag chain; (2) substitution at O-3 of rhamnose A restricts the O-Ags to predominantly helical conformations, (3) substitution at O-3 of rhamnose B has only a slight effect on conformation. The common O-Ag conformations across serotypes identified in this work support the assumption that a quadrivalent vaccine containing serotypes 2a and 3a could provide coverage against S. flexneri serotype 3b and serogroup 5.

scholarly journals Bacteriophage-encoded glucosyltransferase GtrII of Shigella flexneri: membrane topology and identification of critical residues

2005 ◽  
Vol 389 (1) ◽  
pp. 137-143 ◽  
Author(s):  
Adele M. LEHANE ◽  
Haralambos KORRES ◽  
Naresh K. VERMA
Keyword(s):  
Sequence Similarity ◽  
Repeat Unit ◽  
Shigella Flexneri ◽  
Membrane Topology ◽  
Transmembrane Helices ◽  
O Antigen ◽  
Repeat Units ◽  
Related Group ◽  
Critical Residues ◽  
O Antigens

The Shigella flexneri serotypes differ in the nature of their O-antigens. The addition of glucosyl or O-acetyl groups to the common backbone repeat units gives rise to the different serotypes. GtrII glucosylates rhamnose III of the O-antigen repeat unit, thus converting serotype Y (which has no modifications to the basic O-antigen repeat unit) into serotype 2a, the most prevalent serotype. In the present study, the topology of GtrII has been determined. GtrII has nine transmembrane helices, a re-entrant loop and three large periplasmic regions. Four critical residues (Glu40, Phe414, Cys435 and Lys478) were identified in two of the periplasmic regions. Despite the lack of sequence similarity between GtrII and the Gtrs from other serotypes, three of the critical residues identified are conserved in the remaining Gtrs. This is consistent with some degree of mechanistic conservation in this functionally related group of proteins.

scholarly journals Molecular detection of all 34 distinct O-antigen forms of Shigella

2009 ◽  
Vol 58 (1) ◽  
pp. 69-81 ◽  
Author(s):  
Yayue Li ◽  
Boyang Cao ◽  
Bin Liu ◽  
Dan Liu ◽  
Qili Gao ◽  
...  
Keyword(s):  
Shigella Flexneri ◽  
Bacterial Species ◽  
Milk Powder ◽  
Shigella Dysenteriae ◽  
Detection Sensitivity ◽  
Clinical Samples ◽  
Shigella Boydii ◽  
O Antigen ◽  
Probe Concentration ◽  
O Antigens

Shigella is the cause of shigellosis or bacillary dysentery, the occurrence of which is estimated to be 165 million cases per year worldwide, resulting in 1.1 million deaths. Rapid and reliable assays for detecting and identifying Shigella in food, environmental and clinical samples are therefore necessary. Shigella species are traditionally identified by their O antigens. This study developed a DNA microarray targeting O-serotype-specific genes to detect all 34 distinct O-antigen forms of Shigella, including Shigella boydii types 1–18, Shigella dysenteriae types 1–13, Shigella flexneri types 1–5 and 6, and Shigella sonnei. A total of 282 strains were used to test the specificity of the microarray, including 186 Shigella and Escherichia coli representative strains, 86 Shigella clinical isolates and ten strains of other bacterial species that are commonly isolated from food or clinical stool specimens. The oligonucleotide probes were printed on the microarray in concentrations from 1 to 100 μM, and 10 μM proved to be the optimal probe concentration. The detection sensitivity for each serotype was 50 ng genomic DNA or 1 c.f.u. in 25 g milk powder sample following a 6 h enrichment in broth. The microarray is specific, sensitive and reproducible, and, to our knowledge, is the first report of a microarray for serotyping all O-antigen forms of Shigella.

scholarly journals Stable Chromosomal Expression of Shigella flexneri 2a and 3a O-Antigens in the Live Salmonella Oral Vaccine Vector Ty21a

2017 ◽  
Vol 24 (12) ◽  
Author(s):  
Madushini N. Dharmasena ◽  
Manuel Osorio ◽  
Kazuyo Takeda ◽  
Scott Stibitz ◽  
Dennis J. Kopecko
Keyword(s):  
Oral Vaccine ◽  
Shigella Flexneri ◽  
Serum Antibody ◽  
Vaccine Vector ◽  
Shigella Dysenteriae ◽  
Antigenic Determinants ◽  
Content Type ◽  
O Antigen ◽  
Shigella Flexneri 2A ◽  
O Antigens

ABSTRACT We have been exploring the use of the live attenuated Salmonella enterica serovar Typhi Ty21a vaccine strain as a versatile oral vaccine vector for the expression and delivery of multiple foreign antigens, including Shigella O-antigens. In this study, we separately cloned genes necessary for the biosynthesis of the Shigella flexneri serotype 2a and 3a O-antigens, which have been shown to provide broad cross-protection to multiple disease-predominant S. flexneri serotypes. The cloned S. flexneri 2a rfb operon, along with bgt and gtrII, contained on the SfII bacteriophage, was sufficient in Ty21a to express the heterologous S. flexneri 2a O-antigen containing the 3,4 antigenic determinants. Further, this rfb operon, along with gtrA, gtrB, and gtrX contained on the Sfx bacteriophage and oac contained on the Sf6 bacteriophage, was sufficient to express S. flexneri 3a O-antigen containing the 6, 7, and 8 antigenic determinants. Ty21a, with these plasmid-carried or chromosomally inserted genes, demonstrated simultaneous and stable expression of homologous S. Typhi O-antigen plus the heterologous S. flexneri O-antigen. Candidate Ty21a vaccine strains expressing heterologous S. flexneri 2a or 3a lipopolysaccharide (LPS) elicited significant serum antibody responses against both homologous S. Typhi and heterologous Shigella LPS and protected mice against virulent S. flexneri 2a or 3a challenges. These new S. flexneri 2a and 3a O-antigen-expressing Ty21a vaccine strains, together with our previously constructed Ty21a strains expressing Shigella sonnei or Shigella dysenteriae 1 O-antigens, have the potential to be used together for simultaneous protection against the predominant causes of shigellosis worldwide as well as against typhoid fever.

O-antigen structure of Shigella flexneri serotype Yv and effect of the lpt-O gene variation on phosphoethanolamine modification of S. flexneri O-antigens

Glycobiology ◽  
2013 ◽  
Vol 23 (4) ◽  
pp. 475-485 ◽  
Author(s):  
Y. A. Knirel ◽  
R. Lan ◽  
S. N. Senchenkova ◽  
J. Wang ◽  
A. S. Shashkov ◽  
...  
Keyword(s):  
Shigella Flexneri ◽  
Gene Variation ◽  
O Antigen ◽  
O Antigens ◽  
Antigen Structure

scholarly journals Increasing the Affinity of an O‐Antigen Polysaccharide Binding Site in Shigella flexneri Bacteriophage Sf6 Tailspike Protein

2020 ◽  
Vol 26 (32) ◽  
pp. 7263-7273
Author(s):  
Sonja Kunstmann ◽  
Olof Engström ◽  
Marko Wehle ◽  
Göran Widmalm ◽  
Mark Santer ◽  
...  
Keyword(s):  
Binding Site ◽  
Shigella Flexneri ◽  
O Antigen ◽  
Tailspike Protein

Antibody Response to Bacterial Antigens: Characteristics of Antibody Response to Somatic Antigens of Salmonella typhimurium

1970 ◽  
Vol 1 (3) ◽  
pp. 219-225
Author(s):  
Y. Fukazawa ◽  
T. Shinoda ◽  
T. Yomoda ◽  
T. Tsuchiya
Keyword(s):  
Antibody Response ◽  
Salmonella Typhimurium ◽  
Shigella Flexneri ◽  
Immunoglobulin M ◽  
Forming Process ◽  
Igg Antibody ◽  
O Antigen ◽  
Igm Antibody ◽  
O Antigens

The character of the antibody response in the rabbit to Salmonella typhimurium somatic (O) antigen was similar to the response to each of several serotypes of Shigella flexneri O antigens, namely a predominance of production of immunoglobulin M (IgM) antibody. Lipopolysaccharide protein (LPSP) and lipopolysaccharide (LPS) fractions of Salmonella O antigen differed significantly in both quantitative and qualitative aspects of their immunogenicity. LPSP elicited high levels of agglutinins and also induced the production of a significant amount of immunoglobulin G (IgG) antibody at a late period. LPS antigen elicited low levels of agglutinins which were exclusively IgM antibody. These results suggested that the chemical nature of the antigen is one important factor in the determination of the character of the antibody response. Further, it is suggested that the protein moiety of the O antigen complex is a carrier active in allowing induction of early IgM and of late IgG antibodies; in contrast, the lipid moiety may compete with this action of the carrier protein, thereby suppressing IgG antibody in the primary stage of the antibody-forming process.

Mass Spectrometry and Integrated Virus Detection System Characterization of MS2 Bacteriophage

2005 ◽  
Author(s):  
Rabih E. Jabbour ◽  
Deborah Kuzmanovic ◽  
Patrick E. McCubbin ◽  
Ilya Elashvili ◽  
Charles H. Wick
Keyword(s):  
Mass Spectrometry ◽  
Detection System ◽  
Virus Detection ◽  
Ms2 Bacteriophage ◽  
System Characterization

Applying Probe Electrospray Ionization Mass Spectrometry to Cytological Diagnosis: A Preliminary Study by Using Cultured Lung Cancer Cells

2021 ◽  
pp. 1-10
Author(s):  
Yuki Morimoto ◽  
Takeshi Oya ◽  
Mayuko Ichimura-Shimizu ◽  
Minoru Matsumoto ◽  
Hirohisa Ogawa ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Electrospray Ionization ◽  
Squamous Cell ◽  
Cultured Cells ◽  
Sample Pretreatment ◽  
Cell Types ◽  
Diagnostic Tools ◽  
Ionization Mass ◽  
Diagnostic Modality ◽  
Probe Electrospray Ionization

<b><i>Objectives:</i></b> Cytology and histology are 2 indispensable diagnostic tools for cancer diagnosis, which are rapidly increasing in importance with aging populations. We applied mass spectrometry (MS) as a rapid approach for swiftly acquiring nonmorphological information of interested cells. Conventional MS, which primarily rely on promoting ionization by pre-applying a matrix to cells, has the drawback of time-consuming both on data acquisition and analysis. As an emerging method, probe electrospray ionization-MS (PESI-MS) with a dedicated probe is capable to pierce sample and measure specimen in small amounts, either liquid or solid, without the requirement for sample pretreatment. Furthermore, PESI-MS is timesaving compared to the conventional MS. Herein, we investigated the capability of PESI-MS to characterize the cell types derived from the respiratory tract of human tissues. <b><i>Study Design:</i></b> PESI-MS analyses with DPiMS-2020 were performed on various type of cultured cells including 5 lung squamous cell carcinomas, 5 lung adenocarcinomas, 5 small-cell carcinomas, 4 malignant mesotheliomas, and 2 normal controls. <b><i>Results:</i></b> Several characteristic peaks were detected at around m/z 200 and 800 that were common in all samples. As expected, partial least squares-discriminant analysis of PESI-MS data distinguished the cancer cell types from normal control cells. Moreover, distinct clusters divided squamous cell carcinoma from adenocarcinoma. <b><i>Conclusion:</i></b> PESI-MS presented a promising potential as a novel diagnostic modality for swiftly acquiring specific cytological information.

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