Enzyme-Linked Phage Receptor Binding Protein Assays (ELPRA) Enable Identification of Bacillus anthracis Colonies

Bacteriophage receptor binding proteins (RBPs) are employed by viruses to recognize specific surface structures on bacterial host cells. Recombinant RBPs have been utilized for detection of several pathogens, typically as fusions with reporter enzymes or fluorescent proteins. Identification of Bacillus anthracis, the etiological agent of anthrax, can be difficult because of the bacterium’s close relationship with other species of the Bacillus cereussensu lato group. Here, we facilitated the identification of B. anthracis using two implementations of enzyme-linked phage receptor binding protein assays (ELPRA). We developed a single-tube centrifugation assay simplifying the rapid analysis of suspect colonies. A second assay enables identification of suspect colonies from mixed overgrown solid (agar) media derived from the complex matrix soil. Thus, these tests identified vegetative cells of B. anthracis with little processing time and may support or confirm pathogen detection by molecular methods such as polymerase chain reaction.

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Developing Innolysins Against Campylobacter jejuni Using a Novel Prophage Receptor-Binding Protein

Frontiers in Microbiology ◽

10.3389/fmicb.2021.619028 ◽

2021 ◽

Vol 12 ◽

Author(s):

Athina Zampara ◽

Martine C. Holst Sørensen ◽

Yilmaz Emre Gencay ◽

Dennis Grimon ◽

Sebastian Hougaard Kristiansen ◽

...

Keyword(s):

Campylobacter Jejuni ◽

Receptor Binding ◽

Capsular Polysaccharide ◽

Binding Protein ◽

Log Reduction ◽

Cell Counts ◽

Phage Receptor ◽

Receptor Binding Protein

Campylobacter contaminated poultry remains the major cause of foodborne gastroenteritis worldwide, calling for novel antibacterials. We previously developed the concept of Innolysin composed of an endolysin fused to a phage receptor binding protein (RBP) and provided the proof-of-concept that Innolysins exert bactericidal activity against Escherichia coli. Here, we have expanded the Innolysin concept to target Campylobacter jejuni. As no C. jejuni phage RBP had been identified so far, we first showed that the H-fiber originating from a CJIE1-like prophage of C. jejuni CAMSA2147 functions as a novel RBP. By fusing this H-fiber to phage T5 endolysin, we constructed Innolysins targeting C. jejuni (Innolysins Cj). Innolysin Cj1 exerts antibacterial activity against diverse C. jejuni strains after in vitro exposure for 45 min at 20°C, reaching up to 1.30 ± 0.21 log reduction in CAMSA2147 cell counts. Screening of a library of Innolysins Cj composed of distinct endolysins for growth inhibition, allowed us to select Innolysin Cj5 as an additional promising antibacterial candidate. Application of either Innolysin Cj1 or Innolysin Cj5 on chicken skin refrigerated to 5°C and contaminated with C. jejuni CAMSA2147 led to 1.63 ± 0.46 and 1.18 ± 0.10 log reduction of cells, respectively, confirming that Innolysins Cj can kill C. jejuni in situ. The receptor of Innolysins Cj remains to be identified, however, the RBP component (H-fiber) recognizes a novel receptor compared to lytic phages binding to capsular polysaccharide or flagella. Identification of other unexplored Campylobacter phage RBPs may further increase the repertoire of new Innolysins Cj targeting distinct receptors and working as antibacterials against Campylobacter.

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Investigation of the Relationship between Lactococcal Host Cell Wall Polysaccharide Genotype and 936 Phage Receptor Binding Protein Phylogeny

Applied and Environmental Microbiology ◽

10.1128/aem.00653-13 ◽

2013 ◽

Vol 79 (14) ◽

pp. 4385-4392 ◽

Cited By ~ 70

Author(s):

J. Mahony ◽

W. Kot ◽

J. Murphy ◽

S. Ainsworth ◽

H. Neve ◽

...

Keyword(s):

Cell Wall ◽

Host Cell ◽

Receptor Binding ◽

Binding Protein ◽

Cell Wall Polysaccharide ◽

Protein Phylogeny ◽

Host Cell Wall ◽

Phage Receptor ◽

The Relationship ◽

Receptor Binding Protein

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Genome and Proteome of Campylobacter jejuni Bacteriophage NCTC 12673

Applied and Environmental Microbiology ◽

10.1128/aem.05562-11 ◽

2011 ◽

Vol 77 (23) ◽

pp. 8265-8271 ◽

Cited By ~ 42

Author(s):

Andrew M. Kropinski ◽

Denis Arutyunov ◽

Mary Foss ◽

Anna Cunningham ◽

Wen Ding ◽

...

Keyword(s):

Campylobacter Jejuni ◽

Receptor Binding ◽

Pathogen Detection ◽

Phage Genome ◽

Binding Protein ◽

Pcr Amplification ◽

Homing Endonucleases ◽

Bacterial Detection ◽

Content Type ◽

Receptor Binding Protein

ABSTRACTCampylobacter jejunicontinues to be the leading cause of bacterial food-borne illness worldwide, so improvements to current methods used for bacterial detection and disease prevention are needed. We describe here the genome and proteome ofC. jejunibacteriophage NCTC 12673 and the exploitation of its receptor-binding protein for specific bacterial detection. Remarkably, the 135-kbMyoviridaegenome of NCTC 12673 differs greatly from any other proteobacterial phage genome described (includingC. jejuniphages CP220 and CPt10) and instead shows closest homology to the cyanobacterial T4-related myophages. The phage genome contains 172 putative open reading frames, including 12 homing endonucleases, no visible means of packaging, and a putativetrans-splicing intein. The phage DNA appears to be strongly associated with a protein that interfered with PCR amplification and estimation of the phage genome mass by pulsed-field gel electrophoresis. Identification and analyses of the receptor-binding protein (Gp48) revealed features common to theSalmonella entericaP22 phage tailspike protein, including the ability to specifically recognize a host organism. Bacteriophage receptor-binding proteins may offer promising alternatives for use in pathogen detection platforms.

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Phage receptor binding protein-based magnetic enrichment method as an aid for real time PCR detection of foodborne bacteria

The Analyst ◽

10.1039/c3an01100c ◽

2013 ◽

Vol 138 (19) ◽

pp. 5619 ◽

Cited By ~ 32

Author(s):

Somayyeh Poshtiban ◽

Muhammad Afzal Javed ◽

Denis Arutyunov ◽

Amit Singh ◽

Graham Banting ◽

...

Keyword(s):

Real Time ◽

Receptor Binding ◽

Real Time Pcr ◽

Binding Protein ◽

Pcr Detection ◽

Enrichment Method ◽

Foodborne Bacteria ◽

Magnetic Enrichment ◽

Phage Receptor ◽

Receptor Binding Protein

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Faculty Opinions recommendation of IRBIT, inositol 1,4,5-triphosphate (IP3) receptor-binding protein released with IP3, binds Na+/H+ exchanger NHE3 and activates NHE3 activity in response to calcium.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1135829.592914 ◽

2008 ◽

Author(s):

Ines Armando

Keyword(s):

Receptor Binding ◽

Binding Protein ◽

Ip3 Receptor ◽

Receptor Binding Protein

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Host-Pathogen Adhesion as the Basis of Innovative Diagnostics for Emerging Pathogens

Diagnostics ◽

10.3390/diagnostics11071259 ◽

2021 ◽

Vol 11 (7) ◽

pp. 1259

Author(s):

Alex van Belkum ◽

Carina Almeida ◽

Benjamin Bardiaux ◽

Sarah V. Barrass ◽

Sarah J. Butcher ◽

...

Keyword(s):

Infectious Diseases ◽

Receptor Binding ◽

Pathogen Detection ◽

Host Cells ◽

Emerging Pathogens ◽

Emerging Viruses ◽

Ligand Interaction ◽

Novel Structure ◽

Specificity And Sensitivity

Infectious diseases are an existential health threat, potentiated by emerging and re-emerging viruses and increasing bacterial antibiotic resistance. Targeted treatment of infectious diseases requires precision diagnostics, especially in cases where broad-range therapeutics such as antibiotics fail. There is thus an increasing need for new approaches to develop sensitive and specific in vitro diagnostic (IVD) tests. Basic science and translational research are needed to identify key microbial molecules as diagnostic targets, to identify relevant host counterparts, and to use this knowledge in developing or improving IVD. In this regard, an overlooked feature is the capacity of pathogens to adhere specifically to host cells and tissues. The molecular entities relevant for pathogen–surface interaction are the so-called adhesins. Adhesins vary from protein compounds to (poly-)saccharides or lipid structures that interact with eukaryotic host cell matrix molecules and receptors. Such interactions co-define the specificity and sensitivity of a diagnostic test. Currently, adhesin-receptor binding is typically used in the pre-analytical phase of IVD tests, focusing on pathogen enrichment. Further exploration of adhesin–ligand interaction, supported by present high-throughput “omics” technologies, might stimulate a new generation of broadly applicable pathogen detection and characterization tools. This review describes recent results of novel structure-defining technologies allowing for detailed molecular analysis of adhesins, their receptors and complexes. Since the host ligands evolve slowly, the corresponding adhesin interaction is under selective pressure to maintain a constant receptor binding domain. IVD should exploit such conserved binding sites and, in particular, use the human ligand to enrich the pathogen. We provide an inventory of methods based on adhesion factors and pathogen attachment mechanisms, which can also be of relevance to currently emerging pathogens, including SARS-CoV-2, the causative agent of COVID-19.

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