title: Genetic evidence for the involvement of the S-layer protein gene sap and the sporulation genes spo0A, spo0B, and spo0F in Phage AP50c infection of Bacillus anthracis.
abstract: In order to better characterize the Bacillus anthracis typing phage AP50c, we designed a genetic screen to identify its bacterial receptor. Insertions of the transposon mariner or targeted deletions of the structural gene for the S-layer protein Sap and the sporulation genes spo0A, spo0B, and spo0F in B. anthracis Sterne resulted in phage resistance with concomitant defects in phage adsorption and infectivity. Electron microscopy of bacteria incubated with AP50c revealed phage particles associated with the surface of bacilli of the Sterne strain but not with the surfaces of Δsap, Δspo0A, Δspo0B, or Δspo0F mutants. The amount of Sap in the S layer of each of the spo0 mutant strains was substantially reduced compared to that of the parent strain, and incubation of AP50c with purified recombinant Sap led to a substantial reduction in phage activity. Phylogenetic analysis based on whole-genome sequences of B. cereus sensu lato strains revealed several closely related B. cereus and B. thuringiensis strains that carry sap genes with very high similarities to the sap gene of B. anthracis. Complementation of the Δsap mutant in trans with the wild-type B. anthracis sap or the sap gene from either of two different B. cereus strains that are sensitive to AP50c infection restored phage sensitivity, and electron microscopy confirmed attachment of phage particles to the surface of each of the complemented strains. Based on these data, we postulate that Sap is involved in AP50c infectivity, most likely acting as the phage receptor, and that the spo0 genes may regulate synthesis of Sap and/or formation of the S layer.
authors.affiliation: Division of Bacterial, Parasitic, and Allergenic Products, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland, USA.
authors.forename: Roger D
authors.initials: RD
authors.lastname: Plaut
authors.forename: John W
authors.initials: JW
authors.lastname: Beaber
authors.forename: Jason
authors.initials: J
authors.lastname: Zemansky
authors.forename: Ajinder P
authors.initials: AP
authors.lastname: Kaur
authors.forename: Matroner
authors.initials: M
authors.lastname: George
authors.forename: Biswajit
authors.initials: B
authors.lastname: Biswas
authors.forename: Matthew
authors.initials: M
authors.lastname: Henry
authors.forename: Kimberly A
authors.initials: KA
authors.lastname: Bishop-Lilly
authors.forename: Vishwesh
authors.initials: V
authors.lastname: Mokashi
authors.forename: Ryan M
authors.initials: RM
authors.lastname: Hannah
authors.forename: Robert K
authors.initials: RK
authors.lastname: Pope
authors.forename: Timothy D
authors.initials: TD
authors.lastname: Read
authors.forename: Scott
authors.initials: S
authors.lastname: Stibitz
authors.forename: Richard
authors.initials: R
authors.lastname: Calendar
authors.forename: Shanmuga
authors.initials: S
authors.lastname: Sozhamannan
mesh_terms: D017103:Bacillus Phages / Q000235:genetics / Q000502:physiology*; D001408:Bacillus anthracis / Q000821:virology*; D001409:Bacillus cereus / Q000235:genetics; D001413:Bacillus thuringiensis / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D005816:Genetic Complementation Test; D006790:Host-Parasite Interactions; D016254:Mutagenesis, Insertional; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid; D053585:Virus Attachment
chemical_list: D001426:Bacterial Proteins