Characterization of bacteriophages virulent for Clostridium perfringens and identification of phage lytic enzymes as alternatives to antibiotics for potential control of the bacterium

Abstract

There has been a resurgent interest in the use of bacteriophages or their gene products to control bacterial pathogens as alternatives to currently used antibiotics. Clostridium perfringens is a gram-positive, spore-forming anaerobic bacterium that plays a significant role in human foodborne disease as well as nonfoodborne human, animal, and avian diseases. Countries that have complied with the ban on antimicrobial growth promoters in feeds have reported increased incidences of C. perfringens-associated diseases in poultry. To address these issues, new antimicrobial agents, putative lysins encoded by the genomes of bacteriophages, are being identified in our laboratory. Poultry intestinal material, soil, sewage, and poultry processing drainage water were screened for virulent bacteriophages that could lyse C. perfringens and produce clear plaques in spot assays. Bacteriophages were isolated that had long noncontractile tails, members of the family Siphoviridae, and with short noncontractile tails, members of the family Podoviridae. Several bacteriophage genes were identified that encoded N-acetylmuramoyl-l-alanine amidases, lysozyme-endopeptidases, and a zinc carboxypeptidase domain that has not been previously reported in viral genomes. Putative phage lysin genes (ply) were cloned and expressed in Escherichia coli. The recombinant lysins were amidases capable of lysing both parental phage host strains of C. perfringens as well as other strains of the bacterium in spot and turbidity reduction assays, but did not lyse any clostridia beyond the species. Consequently, bacteriophage gene products could eventually be used to target bacterial pathogens, such as C. perfringens via a species-specific strategy, to control animal and human diseases without having deleterious effects on beneficial probiotic bacteria.

Bacteriophages and bacteriophage-derived endolysins as potential therapeutics to combat Gram-positive spore forming bacteria

“…) but also due to negative public perception of chemically treated food and positive perception of phages (Naanwaab et al . ) together with increasing levels of legislation restricting the use of antibiotics (Seal ). The improper prescription or nonprescription sale of antibiotics in some countries, and also poor patient compliance, has contributed to the rapid spread of multidrug resistance in human pathogens.…”

“…; Seelert et al . ) as well as human pathogens Clostridium difficile , Clostridium perfringens (Seal ; Hargreaves and Clokie ). Despite existing as a minor component of the gut commensal microflora, Cl.…”

“…The expression of a Cl. perfringens endolysin (CP25L) within a probiotic Lactobacillus lactis strain presents an interesting alternative to the use of a replicating phage cocktail (Seal ; Gervasi et al . ) and has demonstrated activity against multiple Cl.…”

Non-antibiotic strategies for the control of necrotic enteritis in poultry

“…The biology of bacteriophages (or phage gene products) and utilizing their antibacterial activity has been an interesting issue (Seal, 2013). They are not reported to infect animal and plant cells, hence being potentially safe alternatives to antibiotics.…”

“…They are not reported to infect animal and plant cells, hence being potentially safe alternatives to antibiotics. Specific bacteriophages virulent for C. perfringens (CP) strains can be employed for NE control in chickens and their molecular characterisation and specific bacteriolytic enzymes have recently been conducted (Seal, 2013). Bacterial viruses virulent for C. perfringens have been initially screened by employing filtered samples collected from the intestinal contents of poultry, soil and processing drainage water.…”