title: Peptides panned from a phage-displayed random peptide library are useful for the detection of Bacillus anthracis surrogates B. cereus 4342 and B. anthracis Sterne.
abstract: Recent use of Bacillus anthracis as a bioweapon has highlighted the need for a sensitive monitoring system. Current bacterial detection tests use antibodies as bio-molecular recognition elements which have limitations with regard to time, specificity and sensitivity, creating the need for new and improved cost-effective high-affinity detection probes. In this study, we screened a commercially available bacteriophage-displayed random peptide library using Bacillus cereus 4342 cells as bait to identify peptides that could be used for detection of Bacillus. The method enabled us to identify two 12-amino acid consensus peptide sequences that specifically bind to B. cereus 4342 and B. anthracis Sterne, the nonpathogenic surrogates of B. anthracis strain. The two Bacillus-binding peptides (named BBP-1 and BBP-2) were synthesized with biotin tag to confirm their binding by four independent detection assays. Dot-blot analysis revealed that the peptides bind specifically to B. cereus 4342 and B. anthracis Sterne. Quantitative analysis of this interaction by ELISA and fluorometry demonstrated a detection sensitivity of 10(2) colony forming U/ml (CFU/ml) by both assays. When the peptides were used in combination with Qdots, the sensitivity was enhanced further by enabling detection of even a single bacterium by fluorescence microscopy. Immunoblot analysis and protein sequencing showed that BBP-1 and BBP-2 bound to the S-layer protein of B. anthracis Sterne. Overall, our findings validate the usefulness of synthetic versions of phage-derived peptides in combination with Qdot-liquid nanocrystals as high sensitivity bioprobes for various microbial detection platforms.
authors.affiliation: Section of Cell Biology, Laboratory of Cellular Hematology, Center for Biologics Evaluation and Research, FDA, Bethesda, MD 20892, USA.
authors.forename: Shilpakala
authors.initials: S
authors.lastname: Sainath Rao
authors.forename: Ketha V K
authors.initials: KV
authors.lastname: Mohan
authors.forename: Nga
authors.initials: N
authors.lastname: Nguyen
authors.forename: Bindu
authors.initials: B
authors.lastname: Abraham
authors.forename: Galina
authors.initials: G
authors.lastname: Abdouleva
authors.forename: Pei
authors.initials: P
authors.lastname: Zhang
authors.forename: Chintamani D
authors.initials: CD
authors.lastname: Atreya
mesh_terms: D000595:Amino Acid Sequence; D001408:Bacillus anthracis / Q000737:chemistry / Q000302:isolation & purification*; D001409:Bacillus cereus / Q000737:chemistry / Q000302:isolation & purification*; D001694:Biological Warfare; D001710:Biotin / Q000737:chemistry; D004784:Environmental Monitoring; D004797:Enzyme-Linked Immunosorbent Assay; D008856:Microscopy, Fluorescence; D019151:Peptide Library; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000235:genetics
chemical_list: D019151:Peptide Library; D010455:Peptides; D001710:Biotin