Scientific Publications by FDA Staff

J Bacteriol 2014 Mar;196(6):1143-54

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.

Plaut RD, Beaber JW, Zemansky J, Kaur AP, George M, Biswas B, Henry M, Bishop-Lilly KA, Mokashi V, Hannah RM, Pope RK, Read TD, Stibitz S, Calendar R, Sozhamannan S

In order to better characterize the Bacillus anthracis typing phage AP50c, we designed a genetic screen to identify its bacterial receptor. Mariner transposon insertions 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 Deltasap, Deltaspo0A, Deltaspo0B, or Deltaspo0F mutants. The amount of Sap in the S-layer of each of the spo0 mutant strains was substantially reduced compared to the parent strain, and incubation of AP50c with purified recombinant Sap led to 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 that of B. anthracis. Complementation of the Deltasap 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.