Submitted to: American Society for Microbiology
Publication Type: Abstract Only
Publication Acceptance Date: April 7, 2009
Publication Date: May 17, 2009
Citation: Simmons, I.M., Siragusa, G.R., Donovan, D.M., Seal, B.S. 2009. Comparison of two bacteriophage derived enzymes with lytic activity against strains of Clostridium perfringens. American Society for Microbiology. Technical Abstract: Clostridium perfringens is a Gram-positive anaerobic spore-forming bacterium capable of producing four major toxins which are responsible for disease symptoms and pathogenesis in a variety of animals, humans and poultry. The organism is the third leading cause of food-borne bacterial disease among humans. In chickens C. perfringens is the presumptive etiologic agent of necrotic enteritis. Necrotic enteritis can cause increased mortality in broiler flocks in its acute form and in the sub-clinical form it can lead to decreased digestion and reduced weight gain due to damage of the intestinal mucosa. While it can be controlled by antibiotics, there is increasing pressure to ban growth promoting antimicrobials in poultry feed due to the increasing frequency of antibiotic resistant strains. This ban has yielded an increased incidence of C. perfringens-associated necrotic enteritis in poultry from countries that have curtailed the use of antibiotics in feed. To address this need, we are attempting to identify new antimicrobial agents, putative lytic enzyme genes from the genomes of bacteriophages. Two putative lytic enzyme genes from phiCP39O and phiCP26F were cloned, expressed in E. coli, the resultant proteins purified to homogeneity, and C-termini subjected to protein sequencing. The actual and predicted amino acid sequences of the two recombinant proteins were identical at the C-terminus (cell-wall binding domain), but only 55 per cent similar to each other at the N-terminal catalytic domain. Both recombinant lytic enzymes were able to lyse both parental phage host strains of C. perfringens in turbidity reduction assays. This reduction in turbidity was accompanied by up to a 3 log cfu/ml reduction in viable C. perfringens on BHI (brain heart infusion) agar plates.