Isolation of Salmonella Bacteriophages from Swine Effluent Lagoons

Abstract

Bacteriophages (phages) associated with Salmonella were collected from nine swine manure lagoons in Mississippi. Phages were isolated by an enrichment protocol or directly from effluent. For enrichment, chloroform-treated samples were filtered (0.22 mum) and selectively enriched by adding a cocktail of Salmonella strains in trypticase soy broth. After overnight incubation at 35 degrees C, chloroform was added and samples stored at 5 degrees C. Enriched samples were tested by double agar layer (DAL) plaque assay against individual Salmonella isolates. Phage titers of 2.9 x 10(8) to 2.1 x 10(9) plaque forming units (pfu) per mL were produced, but estimation of phage titers in lagoons was not possible. For direct isolation, effluent was clarified by centrifugation, filtered (0.22 microm), and used in DAL plaque assays to select single-plaque isolates for 15 Salmonella strains. Plaque counts varied among Salmonella strains and lagoons. The most sensitive strain for direct phage recovery was ATCC 13311. Phage titers estimated by direct isolation with ATCC 13311 ranged among lagoons from 12 to 148 pfu per mL. In limited host range tests, 66 isolates recovered by the enrichment protocol produced plaques only on Enteritidis and Typhimurium strains of Salmonella and none produced plaques on lagoon isolates of Citrobacter, Escherichia, Proteus, Providencia, or Serratia. Electron microscopy (EM) showed purified enrichment isolates had Podoviridae morphology (tailless 50-nm icosahedral heads with tail spikes). Electron microscopy of clarified concentrated effluent showed 5.5:1 tailless to tailed phages. The isolated phages have potential as typing reagents, specific indicators, and biocontrol agents of Salmonella.

Phage Therapy To Reduce PreprocessingSalmonellaInfections in Market-Weight Swine

“…We predicted that employing phage therapy to combat reactivating or rapid Salmonella infections associated with transport/lairage would be an ideal use of phage therapy. First, as demonstrated here and by others (19,23), Salmonella-specific virulent phages are readily isolated from various environments. Second, as pigs are not commonly held for over 12 hours during transport and holding, problems of phage resistance development are minimized which, while common in the laboratory, are actually rare under natural conditions (8,13).…”

“…Wild-type phages were isolated using previously described protocols (19,23) with some variations. Wastewater samples were collected from 14 different municipal wastewater treatment plants located throughout Indiana.…”

Temporal flux and spatial dynamics of nutrients, fecal indicators, and zoonotic pathogens in anaerobic swine manure lagoon water

“…2d) and increased competition and predation from other bacteria and microorganisms, including bacteriophages (Callaway et al, 2011). Salmonella-specific bacteriophages have been isolated from water samples collected from this and similar lagoons in the area (McLaughlin et al, 2006), although their temporal distribution and concentrations are unknown. Furthermore, the dynamics of Salmonella population flux in lagoon water may not be fully explained by lagoon factors alone.…”

Bacteriophage PPST1 Isolated from Hospital Wastewater, A Potential Therapeutic Agent Against Drug Resistant Salmonella enterica subsp. enterica serovar Typhi

“…From previous studies, Salmonella bacteriophages were found to be heterogeneous, comprising all of the three families of tailed bacteriophages, including Siphoviridae, Myoviridae and Podoviridae (Demczuk et al, 2004;McLaughlin, 2006;De Lappe et al, 2009). The criteria commonly used for the classification of bacteriophages to a particular group are bacteriophage genetic and morphological characteristics.…”

“…Bacteriophages generally reside in environments that are habitats for their host bacteria (McLaughlin et al, 2006;Sundar et al, 2009). Wastewater from Sappasitthiprasong hospital, Ubon Ratchathani, Thailand was ideal for isolation of bacteriophages in this study since the drug resistant S. Typhi strain SSH1 used as the main host in the present bacteriophage screening is a clinical strain isolated from a patient admitted to the hospital.…”