Bacteriophage therapy to reduce salmonella colonization of broiler chickens

Abstract

Acute enteric infections caused by salmonellas remain a major public health burden worldwide. Poultry, particularly chickens, are known to be the main reservoir for this zoonotic pathogen. Although some progress has been made in reducing Salmonella colonization of broiler chickens by using biosecurity and antimicrobials, it still remains a considerable problem. The use of host-specific bacteriophages as a biocontrol is one possible intervention by which Salmonella colonization could be reduced. A total of 232 Salmonella bacteriophages were isolated from poultry farms, abattoirs, and wastewater in 2004 and 2005. Three phages exhibiting the broadest host ranges against Salmonella enterica serotypes Enteritidis, Hadar, and Typhimurium were characterized further by determining their morphology and lytic activity in vitro. These phages were then administered in antacid suspension to birds experimentally colonized with specific Salmonella host strains. The first phage reduced S. enterica serotype Enteritidis cecal colonization by > or = 4.2 log10 CFU within 24 h compared with controls. Administration of the second phage reduced S. enterica serotype Typhimurium by > or = 2.19 log10 CFU within 24 h. The third bacteriophage was ineffective at reducing S. enterica serotype Hadar colonization. Bacteriophage resistance occurred at a frequency commensurate with the titer of phage being administered, with larger phage titers resulting in a greater proportion of resistant salmonellas. The selection of appropriate bacteriophages and optimization of both the timing and method of phage delivery are key factors in the successful phage-mediated control of salmonellas in broiler chickens.

FIG. 1.

Electron photomicrographs of Salmonella phages φ10 (A), φ25 (B), and φ151 (C). Phages φ10 and φ25 both exhibit icosahedral heads and flexible tails typical of members of the Siphoviridae family. With its icosahedral head and contractile tail, phage φ151 typifies members of the Myoviridae family. The bars represent 250 nm.

FIG. 2.

Graphs A1, B1, and C1 show the optical density readings (at 600 nm) of NB cultures of S. enterica serotypes Enteritidis P125109, Hadar 18, and Typhimurium 4/74, respectively, over a 24-h period. Phages φ151, φ25, and φ10 were separately added to exponential growth phase cultures of S. enterica serotypes Enteritidis, Hadar, and Typhimurium, respectively, at MOIs of 10° (○), 10³ (□), and 10⁶ (⋄). Optical density readings taken from uninfected cultures are also shown (•). Graphs A2, B2, and C2 show the plate counts of bacteriophages (▪) and uninfected (•) and infected (⋄) Salmonella cultures recorded from the same samples (only data for the highest MOIs are presented). All means and standard deviations were calculated by using data from six replicates (optical density) or plate counts (culture) per MOI.

FIG. 3.

Efficacy of high-titer phage treatment in reducing Salmonella counts in the ceca of broiler chickens (trial 2). Broiler chickens colonized by S. enterica serotype Enteritidis P125109 (A), Hadar 18 (B), or Typhimurium 4/74 (C) were treated separately with 11.0 log₁₀ PFU of phage φ151, φ25, or φ10, respectively. Salmonella counts from the ceca of individual control birds (▪) and phage-treated birds (□) are presented as log₁₀ CFU g⁻¹ cecal content.