Characterization of LysPBC4, a novelBacillus cereus-specific endolysin of bacteriophage PBC4

Abstract

Bacillus cereus is a spore-forming, Gram-positive bacterium and is a major food-borne pathogen. A B. cereus-specific bacteriophage PBC4 was isolated from the soil of a stock farm, and its genome was analyzed. PBC4 belongs to the Siphoviridae family and has a genome consisting of 80 647-bp-long double-stranded DNA, including 123 genes and two tRNAs. LysPBC4, the endolysin of PBC4, has an enzymatically active domain (EAD) on its N-terminal region and a putative cell wall-binding domain (CBD) on its C-terminal region, respectively. Although the phage PBC4 showed a very limited host range, LysPBC4 could lyse all of the B. cereus strains tested. However, LysPBC4 did not kill other bacteria such as B. subtilis or Listeria, indicating that the endolysin has specific lytic activity against the B. cereus group species. Furthermore, LysPBC4_CBD fused with enhanced green fluorescent protein (EGFP) could decorate limited strains of B. cereus group, suggesting that the LysPBC4_CBD may be a promising material for specific detection of B. cereus.

Development of a specific fluorescent phage endolysin for in situ detection of Clostridium species associated with cheese spoilage

“…A GFP fusion protein of CPF369_CBD, a putative CBD identified from the genome of C. perfringens ATCC 13124, only bound to C. perfringens cells (Kong and Ryu, ) while a GFP fusion protein of the C‐terminal domain of PlyL, an endolysin encoded by a Bacillus anthracis prophage, bound to Bacillus cereus but not to Bacillus megaterium or Bacillus subtilis cells (Low et al ., ). A fusion of EGFP (enhanced GFP) with PlyG, an endolysin produced by gamma phage, or its truncated C‐terminal domains, specifically recognized B. anthracis cells (Yang et al ., ) while an EGFP fusion protein of the C‐terminal domain of LysPBC4, the endolysin of a B. cereus ‐specific bacteriophage, could only decorate limited strains of B. cereus (Na et al ., ). In the case of non‐spore‐forming bacteria, GFP‐CBDs of Listeria phage endolysins Ply500, Ply118 and PlyP35 only bound to the Listeria genus and showed binding specificity at species and even serovar level (Loessner et al ., ; Schmelcher et al ., ).…”

Genomic characterization of three novel Basilisk-like phages infecting Bacillus anthracis

“…By comparison, the Basilisk phage displayed broad host range infecting B. cereus, B. anthracis, and B. thuringiensis [ 8 ] (Table 2 ). Phage PBC4, in contrast, exhibits pronounced host specificity for B. cereus strain 14,579 [ 9 ].…”

“…In the present study, we sequenced the complete genomes of three novel bacteriophages v_B-Bak1, v_B-Bak6, v_B-Bak10 previously isolated from historical anthrax burial sites in the South Caucasus country of Georgia. Comparative whole genome sequence analysis revealed a close evolutionary relationship between the three newly characterized phages and phage Basilisk, a broad host range B. cereus group phage [ 8 ] as well as phage PBC4, a highly host-restricted B. cereus phage and close relative of Basilisk [ 9 ]. We report here major trends in the molecular evolution of these phages, which we designate as “Basilisk-Like-Phages” (BLPs), and illustrate patterns in their evolution, genomic plasticity and core genome architecture.…”

Characterization of the first double-stranded RNA bacteriophage infecting Pseudomonas aeruginosa

“…We also identified a muramidase, which showed 45% similarity to phi13, from segment S. Muramidase is a phage-encoded lysin that damages bacterial cell walls to release phage particles during the last step of the phage infection cycle2021. Given the emergence of multi-drug resistant bacteria, phage lysin has been suggested to be a potential antimicrobial agent22. However, all the currently tested lysins were cloned from dsDNA phages.…”