Control of Salmonella Gallinarum (Fowl Typhoid) in Poultry with Phage-Based Interventions

Saud Ur Rehman, Purdue University

Abstract

The Pakistan poultry industry has developed into the 11th largest poultry industry in the world and poultry products provide high-quality and affordable protein sources to communities throughout the country. However, Salmonella Gallinarum, the etiological agent for fowl typhoid, is endemic in Pakistan with infections leading to high mortality and substantial economic loss. Currently, Salmonella Gallinarum infections in Pakistan poultry are controlled with antibiotics. The continued emergence of antibiotic resistance, however, has led to global initiatives to reduce the use of antibiotics in both human and veterinary medicine. Concurrently, the Pakistan government recently introduced new national policies that limit the use of antibiotics for performance in livestock and poultry production. As such, controlling bacterial infections in poultry without increasing the likelihood of antibiotic use could ensure the sustainability of Pakistan poultry production without posing risks to public health. Toward this end, we hypothesized that Salmonella Gallinarum infections in chickens could be prevented or otherwise controlled through the use of phages. To test this hypothesis, wastewater samples were collected from Lahore, Pakistan and different cities of Indiana, US and processed to isolate bacteriophages. The phages were characterized in terms of morphology, host spectra, lytic capacity, genomic sequencing, and survivability in different environments. Transmission electron microscopy showed these phages belonged to myoviridae (n = 5) and podoviridae (n = 1) families. Spectrum analysis revealed that each phage lysed at least 8 out of 10 different strains of Salmonella Gallinarum and significantly reduced (P < 0.05) Salmonella Gallinarum when co-cultured in liquid medium with the bacterium. Stability of the phages was tested in simulated gastric fluid (SGF; pH = 2.5) and simulated intestinal fluid (SIF; pH~6.8). Results showed that phage concentrations were reduced to undetectable levels when exposed to SGF for more than 5 minutes. However, exposure to SIF did not result in appreciable reductions in phage concentrations. To mitigate potential effects of gastric environments, phages were encapsulated using a sodium alginate-based method. In contrast to unprotected phages, encapsulated phages remained viable (~100%) after 30 minutes exposure to SGF. Additionally, encapsulation efficiencies ranged between 90-99%. Encapsulated phages were sequentially incubated in SGF (30 minutes) and SIF (120 minutes) to determine the rate of release of the phages from capsules. All phages were released from capsules after 60 minutes of exposure to SIF. To determine if the phages effectively controlled Salmonella Gallinarum infections in chickens, 100, day-old Jumbo Cornish Rock Cross birds were randomly assigned to one of four treatments: 1) Control 1 (bacterial challenge, no phage treatment); 2) Control 2 (no phage or bacterial challenge); 3) challenged with Salmonella Gallinarum and treated with unprotected phages; and 4) challenged with Salmonella Gallinarum and treated with encapsulated phages. At 7 d of age, chicks receiving the bacterial challenge were administered 5 X 106 CFU (500 μL) of Salmonella Gallinarum. For birds in phage treatment groups, the phages were administered (500 uL; 5 X 108 PFU/mL or g) at 0, 12, and 24 hours post-challenge. Six birds from each group were euthanized at 1, 2, and 4 days post-challenge (dpc) and cecal Salmonella Gallinarum concentrations were quantified. At 1 dpc, birds treated with unprotected and encapsulated phages had significantly lower (P < 0.05) SalmonellaGallinarum concentrations (4.36 ± 0.20 and 5.05 ± 0.22 log CFU/g, respectively) than those found in untreated birds (5.71 ± 0.13).

Degree

M.Sc.

Advisors

Ebner, Purdue University.

Subject Area

Animal sciences|Pathology

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