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Dissecting virulence of E. coli O157:H7 using genome alignments

Dissecting virulence of E. coli O157:H7 using genome alignments. The first E. coli genome sequenced was the non-pathogenic E. coli K-12 genome MG1655. - determination of the complete E. coli sequence required almost 6 years

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Dissecting virulence of E. coli O157:H7 using genome alignments

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  1. Dissecting virulence of E. coli O157:H7 using genome alignments

  2. The first E. coli genome sequenced was the non-pathogenic E. coli K-12 genome MG1655 -determination of the complete E. coli sequence required almost 6 years -E. coli is the preferred model in biochemical genetics, molecular biology, and biotechnology and its genomic characterization will undoubtedly further research toward a more complete understanding of this important experimental, medical, and industrial organism (Blattner et al. Science 1997)

  3. The first pathogenic E. coli genome sequence was enterohaemorrhagic (EHEC) Escherichia coli O157:H7 strain 933 EDL -In 1982 Escherichia coli O157:H7recognized as a pathogen for human disease -Also known as EDL933 from the Michigan outbreak in 1982 from ground beef -shiga toxin producing (STEC) Epidemiological Statistics -47 sickened individuals, no deaths (Perna et al. Nature 2001)

  4. Comparison of E. coli K-12 vs. E. coli O157:H7 The Good vs. the Bad 1,000 extra genes Where are the differences? ~4,200 genes 4,639,675 bp ~5,200 genes 5,528,423bp

  5. E. coli O157:H7 EDL933 (pathogenic) vs. E. coli K-12 MG1655 (non-pathogenic) -Areas in red are only found in E. coli K12 specific -Areas in orange are E. coli O157:H7 specific -Areas in blue are conserved in both -Regions that are unique to one genome are called “islands” EDL933 islands K-12 islands Backbone

  6. A 3rd type of E. coli Genome sequenced was the uropathogenic Escherichia coli strain CFT073 -Uropathogenic strains of Escherichia coli (UPEC) are the most common cause of non-hospital-acquired urinary tract infections, responsible for 70-90% of the 7 million cases of acute cystitis and 250,000 cases of pyelonephritis reported annually in the United States. -the highly virulent UPEC strain CFT073, isolated from the blood of a woman with acute pyelonephritis. -Represented the 3rd type of E. coli genome sequenced (Welch et al. 2002, PNAS)

  7. Three E. coli Genomes comparison of gene content: the good the bad and the ugly KO=396 Kb Ki=329 Kb Oi=1.248 Mb KOC = 3.711 Mb KC= 144 Kb OC=170 kb Ci = 1.125 Mb K-12 (Good) O157:H7 (Bad) i=islands CFT073 (Ugly)

  8. Three E. coli Genomes comparison (cont.) -difference in disease potential between O157:H7 and CFT073 is reflected in the absence of genes for type III secretion system or phage- and plasmid-encoded toxins -CFT073 genome is particularly rich in genes that encode potential fimbrial adhesins, autotransporters, iron-sequestration systems, and phase-switch recombinases -Comparisons indicate that extraintestinal pathogenic E. coli arose independently from multiple clonal lineages (Welch et al. 2002, PNAS)

  9. The completion of the 2ndE. coli O157:H7 (EHEC) sequence strain Sakai • -In July 1996, an outbreak of Escherichia coli O157:H7 infection occurred among schoolchildren in Sakai City, Osaka, Japan. • Epidemiological Statistics • 8,938 schoolchildren sickened, 3 deaths • We are starting to ask-What genomic differences determine differences in virulence, epidemiology, and fatality? (Hayashi et al. DNA Res 2001)

  10. In 2006 E. coli O157:H7 outbreakfrom bagged spinach (from CDC) -multistate outbreak Epidemiological Statistics 205 people sickened, 3 deaths -Produce associated outbreak strains caused higher incidence of hemolytic-uremic syndrome (HUS) (Manning et al. PNAS 2008) -genome alignments can be used to find variations

  11. Epidemiology of E. coli O157:H7 Outbreaks, United States, 1982-2002 Median size of E. coli O157:H7 outbreaks by year E. coli O157:H7 outbreaks by year 1982-2002 (Rangel et al. 2005)

  12. Epidemiology of E. coli O157:H7 Outbreaks, United States, 1982-2002 (cont.) Vehicles of foodborne E. coli O157:H7 outbreaks by year Transmission routes of E. coli O157:H7 outbreaks by year (Rangel et al. 2005)

  13. In the most severe cases, E. coli O157:H7 causes hemolytic uremic syndrome (HUS) and death Hemolytic uremic syndrome (HUS) and case-fatality rate per 100 outbreak-related illnesses (Rangel et al. 2005)

  14. Do genomes change over time? Yes, the genome sequence represents a snapshot at the time the strain was sequenced. Genomes may vary ~15% or more over time Courtesy of fbi.gov For example E. coli O157:H7 strain EDL933 originally had two copies of a TAI pathogenicity island In the lab, it has lost one of the tellurite resistance-and adherence-conferring island TAI pathogenicity islands (unpublished data)

  15. Currently there are 15 E. coli O157:H7 Genomes sequenced, we will have you focus on three that are all in the Enteropathogen Resource Integration Center (ERIC) database (www.ericbrc.org) The three strains you will focus on are: Escherichia coli EDL933 (EHEC) [ground beef 1982 outbreak] Escherichia coli Sakai (EHEC) also called RIMD [radish sprouts 1996 outbreak] Escherichia coli EC4042 (EHEC) [fresh bagged spinach 2006]

  16. Genome comparisons provide us to start to understand differences that have occurred over time and geographical differences, and also investigate variations in the genomes that may have consequences for virulence differences Courtesy of USGS

  17. Comparative genomics using the tool Mauve: Multiple Genome Aligner • Able to identify conserved regions of multiple genomes even in the presence of rearrangements • (Darling et al. 2004)

  18. Your individual projects Investigate a known or putative virulence factor and determine if it is conserved, and also if it is in the same place in the genomes of the three E. coli O157:H7 strains. Types of virulence factors: Toxins, attachment and effacement, adhesion, effectors, defenses against host barriers Also using the Mauve alignment, look for islands unique to 1 out of 3 and also 2 of the 3 of the E. coli O157:H7 strains

  19. Please choose one and sign up on the list and provide your email address ABH-0027064 stx1B ABH-0027065 stx1A ABH-0025275 stx2A ABH-0025276 stx2B ABH-0028752 eae ABH-0028754 tir ABH-0024965 ureD ABH-0026806 tccP ABH-0025423 iha ABH-0024997 terC ABH-0028614 lpfC ABH-0026805 espJ For more putative and known virulence factors (320 identified) see attached excel spreadsheet called List of Virulence factors in Ecoli O157:H7 strain EDL933

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