Report on the 9th International

Symposium on Shiga Toxin

producing E. coli infections

(VTEC 2015):

main breakthroughs

and the way forward

Flemming Scheutz

WHO Collaborating Centre forReference and Research on

Escherichia and Klebsiella

Foodborne bacterial infections

Microbiology & Infection controlSTATENS SERUM INSTITUT

Flemming Scheutz, Copenhagen

Reinhard Würzner, Innsbruck

Summary of highlights of all oral sessions

”Wisdom entereth not into a malicious mind,

and science without conscience is but the

ruin of the soul”

François Rabelais, 1532

Once upon a time….

7

2324 E. coli genomes – Panseq SNP phylogeny (ML)

O157

O12

1

O11

3

O145

O2

6

Courtesy of Chad Laing

Once upon a time….

These days:Outbreaks:

• Watercress

Risk factors:

• Swimming & camping (O26; 47%)

• Contact with ruminants (O103; 36%)

• Contact with another person with diarrhoea (Children)

• Visiting farms (O103), (Children)

• Steaks and ground beef (Children)

• Travel to Africa (Turkey and Marocco) (Adults)

Host factors:

• Antaacids (Adults)

• Cardiovascular disease (Adults)

• Gastrointestinal diseases (Adults)

Foods

Hispanic ethnicity

No significant risk factors for O111; 17%

Hamburgers =

ground beef

I.H.M. Friesema et al. NL

Ellyn Marder et al. USA

Non-O157s now share the scene with O157

from cases of HUS

O26:H11 in Italy (stx2a highly virulent)

O80:H2 in France (stx2a, stx2a2d, stx2c2d, stx2d, eae ξ)

O104:H4 in the republic of Georgia

O73:H18 in Spain

O91:H21 in Spain

O59:H19 EAEC in Argentina

O174:H8, H21 and H28 in Argentina

Lineages

O157:H7

• some lineages are quite domesticated;

others are ”tourists” (USA, Australia,

Argentina)

• Some are host specific

O91:H21, H14, H10 & H9

• H type predictive of virulence profile and

MLST type

O103:H25 (Norway)

• Two main, one of which human and bovine

Reservoir &

Supershedder

s

Cattle reservoir

• Seasonal patterns due extrinsic factors

• The decline in human O157 infections mirrors

the decrease of findings in beef

• SNPs analyses did not support the genetic

segregation between super-shedder or low-

shedder isolates

• Reduced immune response in super shedders

Do the bacteriae reduce this immune response or is

it host re(gu)lated?

Allelic signatures can be used to classify VTEC strains into

groups segregating with the serogroups and discriminate

subpopulations within the single groups

• gnd – the ‘passive hitch hiker’

• locus for enterocyte effacement (LEE)

• PAIs OI-122 and OI-57

• λ-like phages contain genetic factors capable of

controlling STEC virulence including the LEE

pathogenicity island and genes associated with adhesion

• Only a fraction (60% 4-6 hours and 80% after 24 hours)

of the cells induces Stx phages

• Population bistability in phage induction could be a

mechanism positively selected to prevent the extinction

of the STEC lysogenic population

• Bistability can be modulated by environmental

conditions eg under stress conditions or different growth

stages using the regulator RpoS

Bacteriophages and their hosts

Bacteriophages

Stx2f bacteriophage originating in pigeons

may have gone into aEPEC via other lower

pathogenic E. coli types:

O26:H11

O55:H9

O80:H2

OI-122, pO157 plasmid

Courtesy of Stefano Morabito; One Health VTEC-HUS Symposium; 10th December 2014

H10

Next generation sequencing

NGSor

Whole genome sequencing

WGS

O157 can be given an SNP address which can

be correlated with

• Geographical region

• Reservoir

Epidemiology

Acknowledgments

20 Application of WGS to outbreak investigation of gastrointestinal pathogens

• 1,075 strains of STEC O157 from clinical and animal isolates from

England, Northern Ireland, Wales and Scotland collected from 1985 to

2014

Application of WGS to outbreak investigation of gastrointestinal pathogens

Maximum clade credibility

tree of 530 ∆25 SNP

representatives of O157:H7

= Family Tree of O157:H7

using ∆25 SNP addresses

SNP = Single Nucleotide

Polymorphism

Tim Dallman et al. Microbial Genomics 2015

http://benfry.com/zipdecode/

Application of WGS to outbreak investigation of gastrointestinal pathogens

175 years

~1840

150 years

400 years

The ancestral O157:H7 state is stx2c

(lineage II)

Early rapid diversification

Shiga toxin complement and insertion

sites are relatively stable

Stx2a is relatively recent and has been

acquired ~60 years ago on multiple

occasions

Courtesy of Tim Dallman

YEARS

Courtesy of Tim Dallman

O157:H7 has become more pathogenic to humans

over its evolution

Strong association with stx2a and severe disease

YEARS

25 Application of WGS to outbreak investigation of gastrointestinal pathogens

Uneven Diversity of O157

54 ∆250 Clusters

All 3 lineages have one ∆250 cluster

that comprises > 50% of isolates of that

lineageThese 3 ∆250 clusters contains 84% of UK

Cattle isolates

∆250 clusters with <5 isolates

associated with foreign travel or

imported food

Supports Global Dispersion then Regional Expansion

Courtesy of Tim Dallman

26 Courtesy of Tim Dallman

The implicated O157

strain with the SNP

address 18.35.397.765

had a strong NW

geographical signal

including some possible

environmental exposures

18.yy.3xx.zzz has a strong

individual signal

including some possible

VTEC symposium

exposures

Heat Resistance

O69:H11

ONT:H25

O11:H30

Will survive 60 ◦C and associated with a clpK

encoded heat resistance gene

Prevalence of VTEC in healthy humans

including food handlers or workers in daycare

centers during 2010-2012 in Japan

Healthy carriers Diarrhoeal cases

84.2 2.1

/100,000 /100,000

Tomoko Morita-Ishihara et al. IASR 2015

The way ahead

• Our perception of ”rare”

• Definition of case – control studies

• Clarify legal aspects of deposition of WGS in the public

domain

• Extract the different gene cocktails associated with

(severe) disease (WGS)

• Relate WGS data to the biology

• Identify host factors including ”WHO’S THE BOSS”?

The way ahead

• Early detection – early treatment

• Immunizations?

• Acute eHUS: Increase volume!

• Identify further virulence & host factors to find new targets

• Clarify the role of complement – are there indications for

Eculizumab?

• NO antibiotics in the preHUS until we have a safe

candidate

• Consider antibiotics in eHUS to enforce clearance

ACKNOWLEDGEMENTS

• Tim Dallman, Public Health England

• Chad Laing; Public Health Agency of

Canada

• Stefano Morabito; Istituto Superiore di

Sanità, Rome

New paradigms?

Try to read this:

TH15 M3554G3 53RV35 T0 PR0V3 H0W OUR

M1ND5 C4N D0 4M4Z1NG TH1NG5!

1MPR3SS1V3 TH1NG5! 1N TH3 B3G1NN1NG 1T

W4S H4RD BUT N0W, 0N TH15 L1N3 Y0UR

M1ND 1S R34D1NG 1T 4UT0M4T1C4LLY W1TH

PR4T1C4LLY N0 TH1NK1NG 1NV0LV3D

R1GHT? B3 V3RY PR0UD ! Y0U D35ERVE 4 P4T

0N TH3 B4CK!