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Escherichia coli O157:H7 and Shiga toxin-converting bacteriophage Edward G. Dudley, Ph.D Department of Food Science [email protected] Escherichia coli O157:H7 history and food safety issues Foodborne illness affects 48 million annually http://wwwnc.cdc.gov/eid/article/17/1/pdfs/p2-1101.pdf Escherichia coli O157:H7 is a recently discovered foodborne pathogen E. coli are first characterized by serotyping • O-antigen – Somatic antigen – >170 known • H-antigen – Flagellar antigen – >60 known • E. coli O157:H7 5 Images from Dr. Erika A. Taylor’s website Transmission of O157:H7 to humans usually starts with cattle Nougayrède et al., 2003 Control of E. coli O157:H7 in food supply starts at processing plant Beef represents ~50% of foodborne cases of disease Evolution of one of the defining virulence factors of E. coli O157:H7 Escherichia coli evolves via genomic insertions and deletions “horizontal gene transfer” <30% of genes are shared in all Escherichia coli strains Welch R A et al. PNAS 2002;99:17020-17024 ©2002 by National Academy of Sciences Temperate phage are drivers of most genomic diversity in Escherichia coli diagram from Dr. Ken Todar Many virulence factors are phage encoded Foodborne pathogens: Clostridium botulinum Staphylococcus aureus Vibrio cholerae Salmonella enterica Shigella flexneri Other Escherichia coli Escherichia coli O157:H7 carry an active phage that encodes a toxin Recombination Regulation PR’ Stx2A/B cI = repressor protein Lysis Capsid/tail fiber Plunkett III, G., D. J. Rose, T. J. Durfee, and F. R. Blattner. 1999. Sequence of Shiga toxin 2 phage 933W from Escherichia coli O157: H7: Shiga toxin as a phage late-gene product. J. Bacteriol. 181:17671778. DNA damaging agents induce phage and increase toxin production Ciprofloxacin (DNA gyrase) 14 Shiga toxin blocks protein synthesis by targeting 28S rRNA One “A” subunit (319 amino acids) http://journal.frontiersin.org/Journal/10.3389/fcimb.2012.00081/full AB5 toxin Five “B” subunits (91 amino acids) Do other organisms change amount of toxin produced? Do gut organisms affect virulence of Escherichia coli O157:H7? Experimental setup E. coli O157:H7 strain PA2 + Commensal (non-pathogenic) Plate counts ELISA for toxin production Shiga toxin levels affected by other intestinal E. coli strains Germ free mouse experiments Control Colonize with E. coli C600 Day -7 Test Plate feces Sacrifice 5 mice Inoculate with E. coli O157:H7 Day 0 Day 1 Day 6 Increased virulence of E. coli O157:H7 in presence of non-pathogenic E. coli Acute renal tubular necrosis in mice colonized by PA2 with or without C600 4 p=0.0077 ATN score 3 2 1 0 C600 PA2 PA2+ C600 Growing threats to the food supply The Shiga toxin phage can mobilize to other E. coli, creating new pathogens enteroaggregative Escherichia coli • Serotype O104:H4 • Ill: 4,321. HUS: ~30% of cases. Dead: 50 Several “new” O-groups are of increasing concern Year Serogroup Location Food/event # ill 2014 O111 Minnesota Cabbage (Applebees and Yard House restaurants) 13 O121 5 States Sprouts (Jimmy Johns, Pita Pit) 19 2013 O121 9 States (including PA) Farm Rich Products frozen snacks 35 2012 O145 5 States Unknown 18 2011 O26 OH, PA, AL Sprouts (Jimmy Johns) 29 2010 O145 5 States (including PA) Shredded romaine lettuce 33 The “Big Six”: Shiga toxin E. coli of serogroups O26, O45, O103, O111, O121, O145