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. 2011 Jan;1(1):15-24.
doi: 10.4161/bact.1.1.14175.

Naturally resident and exogenously applied T4-like and T5-like bacteriophages can reduce Escherichia coli O157:H7 levels in sheep guts

Raul R Raya  1 , Rebecca A Oot, Ben Moore-Maley, Serena Wieland, Todd R Callaway, Elizabeth M Kutter, Andrew D Brabban
Affiliations

Naturally resident and exogenously applied T4-like and T5-like bacteriophages can reduce Escherichia coli O157:H7 levels in sheep guts

Raul R Raya et al. Bacteriophage. 2011 Jan.

Abstract

In preparing sheep for an in vivo Escherichia coli O157:H7 eradication trial, we found that 20/39 members of a single flock were naturally colonized by O157:H7-infecting phages. Characterization showed these were all one phage type (subsequently named CEV2) infecting 15/16 O157:H7, 7/72 ECOR and common lab strains. Further characterization by PFGE (genome∼120 kb), restriction enzyme digest (DNA appears unmodified), receptor studies (FhuA but not TonB is required for infection) and sequencing (>95% nucleotide identity) showed it is a close relative of the classically studied coliphage T5. Unlike T5, CEV2 infects O157:H7 in vitro, both aerobically and anaerobically, rapidly adsorbing and killing, but resistant mutants regrew within 24 h. When used together with T4-like CEV1 (MOI ∼2 per phage), bacterial killing was longer lasting. CEV2 did not reproduce when co-infecting the same cell as CEV1, presumably succumbing to CEV1's ability to shut off transcription of cytosine-containing DNA. In vivo sheep trials to remove resident O157:H7 showed that a cocktail of CEV2 and CEV1 (∼10(11) total PFU) applied once orally was more effective (>99.9% reduction) than CEV1 alone (∼99%) compared to the untreated phage-free control. Those sheep naturally carrying CEV2, receiving no additional phage treatment, had the lowest O157:H7 levels (∼99.99% reduction). These data suggest that phage cocktails are more effective than individual phage in removing O157:H7 that have taken residence if the phage work in concert with one another and that naturally resident O157:H7-infecting phages may prevent O157:H7 gut colonization and be one explanation for the transient O157:H7 colonization in ruminants.

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Figures

Figure 1
Figure 1
Electron micrograph of bacteriophage CEV2.
Figure 2
Figure 2
Pulsed-field gel electrophoresis (PFGE) of undigested and restriction enzyme digested bacteriophage CEV2 DNA. (A) Lane A SmaI:CEV2; Lane B EcorI:CEV2; Lane C XhoI:CEV2; Lane D TESC Lab Phage PFGE Ladder consisting of PEV3 [290 kb], T4 [175 kb], T5 [121.9 kb] and PEV2 [72.7 kb] and 816a [42.7 kb]; Lane E NEB HindIII:λ ladder [23.1, 9.4, 6.6, 4.4 kb]; Lane F undigested CEV2; Lane G EcorV:CEV2; Lane H, PvuII:CEV2; Lane I HindIII:CEV2. (B) Lane A HhaI:CEV2; Lane B HaeIII:CEV2; Lane C McrBC:CEV2; Lane D BamHI:CEV2; Lane E TESC Lab Phage PFGE Ladder consisting of PEV3 [290 kb], T4 [175 kb], T5 [120 kb] and PEV2 [70 kb] and 816a [39 kb]; Lane F NEB HindIII:λ ladder [23.1, 9.4, 6.6, 4.4 kb]; Lane G undigested CEV2.
Figure 3
Figure 3
High MOI CEV2 infections of E. coli O157:H7 NCTC 12900 growing either aerobically (A, MOI∼5.3) or anaerobically (B, MOI∼8.3) in TSB. Infection graphs shown are representative of at least three replicates. Bacterial survivors CFU mL−1 (▵), OD600 nm (○) and phage PFU mL−1 after the addition of chloroform (□).
Figure 4
Figure 4
High MOI (∼3.9) co-infection of E. coli O157:H7 NCTC 12900 growing anaerobically in TSB with phages CEV1 (∼2.06) and CEV2 (∼1.88). Bacterial survivors (▵), OD600 nm (○) and phage titers, CEV1 (solid line) and CEV2 (dashed line) after the addition of chloroform (□).
Figure 5
Figure 5
The use of O157:H7-infecting phage CEV1 or a cocktail of CEV1+CEV2 as a pre-slaughter treatment to remove resident E. coli O157:H7 EDL 933 from the intestines of ruminants (sheep). Group 1 (white): Control Group, O157:H7-infecting phage free and receiving no phage treatment. Group 2 (gray): Cocktail (CEV1+CEV2). Treatment Group, O157:H7-infecting phage free and treated with CEV1 and CEV2. Group 3 (black): Naturally Resident CEV2 Phage Group: Sheep in which CEV2 was naturally present. Error bars indicate standard deviations.
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References

    1. Bell BP, Goldoft M, Griffin PM, Davis MA, Gordon DC, Tarr PI, et al. A multistate outbreak of Escherichia coli O157:H7-associated bloody diarrhea and hemolytic uremic syndrome from hamburgers. The Washington experience. JAMA. 1994;272:1349–1353. doi: 10.1001/jama.1994.03520170059036. - DOI - PubMed
    1. Brabban AD, Nelson DA, Kutter E, Edrington TS, Callaway TR. Approaches to controlling Escherichia coli O157:H7, a food-borne pathogen and an emerging environmental hazard. Environmental Practice. 2004;6:208–229. doi: 10.1017/S1466046604000365. - DOI
    1. Nataro JP, Kaper JB. Diarrheagenic Escherichia coli. Clinical Microbiol Rev. 1998;11:142–201. - PMC - PubMed
    1. Mead PS, Griffin PM. Escherichia coli O157: H7. Lancet. 1998;352:1207–1212. doi: 10.1016/S0140-6736(98)01267-7. - DOI - PubMed
    1. Centers for Disease Control and Prevention (CDC), author Ongoing multistate outbreak of Escherichia coli serotype O157:H7 infections associated with consumption of fresh spinach—United States, September 2006. MMWR Morb Mortal Wkly Rep. 2006;55:1045–1046. - PubMed

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