Download Whole Genome Sequencing and foodborne pathogens: a UK

ICMSF WGS Meeting, CCFH 47- Boston, USA – 12 November 2015
Whole Genome Sequencing
and foodborne pathogens: a
UK perspective
Paul Cook
Head of Microbiological Risk Assessment
Science, Evidence and Research Division
[email protected]
Scope of presentation
• WGS trends
• Recommendations from expert workshops
• Salmonella testing by Public health
•
•
England
Examples where WGS has contributed to
outbreak investigations
Other areas where WGS can have an
impact
Trends - rise of the genomes
WGS –PubMed 3 Nov 15
1.
Wetterstrand KA. DNA Sequencing Costs: Data from the NHGRI Large-Scale Genome Sequencing Program Available at:
www.genome.gov/sequencingcosts. Accessed 18 April 2015.
WGS technology is getting
smaller
Converging technologies – can we get closer to the sampling point?
A reference bacterial genome dataset generated on the MinION™
portable single-molecule nanopore sequencer. Quick et al. GigaScience
2014, 3:22 http://www.gigasciencejournal.com/content/3/1/22
Expert workshop 2012
Workshop organised by Food Standards
Agency (FSA), Biotechnology and
Biological Sciences Research Council
(BBSRC), Public Health England (PHE),
UK Advisory Committee on the
Microbiological Safety of Food (ACMSF)
Looked at potential for this approach
to support FSA objectives in tackling
Foodborne illness
Workshop report:
http://www.food.gov.uk/multimedia/pdfs/committee/acm-1092a.pdf
Expert workshop 2012
• WGS capable of providing a significant benefit to
outbreak investigations – timing, specificity
• Technical, quality, logistical and training issues
need to be addressed but should not delay roll-out
of the technology
• Need well-curated, inter-operable and quality
controlled databases containing microbial
sequences
• Need to sequence historical isolates to
help populate the databases – and the
tools to interpret.
EFSA Scientific Colloquium 20 Report
Recommended a need for:
•
•
•
•
•
•
International collaboration to co-ordinate and
harmonise application of WGS
Educational interdisciplinary programmes on WGS for
epidemiologists, bioinformaticians, microbiologists etc
Harmonization of terminology and data reporting in
plain language
Software and data analysis tools that are transparent
and easy to use in WGS data processing
The legal consequences of large-scale application of
WGS should be reviewed
WGS databases for public health - with appropriate
curation standards
June 2014 http://www.efsa.europa.eu/en/supporting/pub/743e
Global Microbial Identifier (GMI)
•
•
•
Built from initial group of 25 participants to current involvement of
around c200 experts from over 30 countries, including clinical-, food-,
and public health microbiologists and virologists, bio-informaticians,
epidemiologists, funding agencies, data hosting systems, and policy
makers from academia, public health, industry, governments.
Steering Committee involving Danish Technical Univ, US FDA, NCBI,
WHO, FAO, FSA, PHE, RIVM, OIE, Thailand Ministry of Public Health,
Chinese National Inst, for Communicable Disease Control and
Prevention
5 active working groups: – Work group 1: Political challenges, outreach, building a global network and
funding
– Work group 2: Repository and storage of sequence and meta-data
– Work group 3: Analytical approaches
– Work group 4: Ring trials and quality assurance
– Work group 5: Pilot projects
– More information: www.globalmicrobialidentifier.org
Whole Genome Sequencing at PHE
Financial investment, machines, robotics, LIMS, data and bioinformatics
workflows, data storage, staffing, training, methodological development –
sample prep, library prep, validation of processes, accreditation ISO15189
2 HiSeq 2500 high-throughput machines
2 MiSeq machines
+
NextSeq machine
Capacity < 3,000 genomes per week
PHE Detection of Salmonella
outbreaks
• Use an ‘exceedance’ above what we would
•
•
•
expect to see as background, before an
outbreak investigation is triggered – based on
serotype and phage type
MLVA, PFGE performed for greater strain
discrimination – but not on every isolate
The more common a serotype is, such as S.
Enteritidis or Typhimurium, the harder it is to
spot an outbreak
Using SNP analysis to detect clusters and
outbreaks
PHE WGS Salmonella workflow
Isolate received by reference lab
undergoes automated DNA extraction
WGS by Genomics Sequencing Unit
GBRU receives FASTQs
Bioinformatic pipelines for K-mer ID
MLST used to assign serotype
Based on serotype
strain goes to
automated SNP
analysis (SNPDB)
for genetic relatedness
and cluster detection
Triggers epidemiological investigation
Report to referring lab
SE PT14b outbreak 198 of 287 (69%) cases could be plausibly linked to
eggs supplied by one company
Two examples of recent outbreaks
in the UK where WGS has been
used
Salmonella Enteritidis and eggs
E.coli O157 and watercress
WORKING OUT WHAT
HAPPENED
Microbiological
Environmental/
Food Chain
Epidemiological
Source: Sarah O’Brien (Univ Liverpool)
= Cause!
Current challenges for incident risk
assessment and management
• Alignment - having the right evidence
•
•
•
from the right place and at the right time
History - event has happened but
environmental evidence cannot be found
Interpretation – differing opinions seeking an independent view
“Weight of evidence” to support decision
making
WGS can help with some of these but not all
Salmonella
Enteritidis
phage type
14b: National
outbreak MaySept 2014
International outbreak of
Salmonella Enteritidis PT14b Summer 2014
• In England an outbreak of
Salmonella Enteritidis PT
14b in a hospital in
Birmingham with single
MLVA profile
• Also noticed small pockets
of increase in PT14b
around England with same
MLVA or SLV – individual
point source outbreaks?
SE PT14b outbreak .198 of 287 (69%) cases could be plausibly linked to
eggs supplied by one company
Inns et al. EuroSurveill.2015;20(16):pii=21098 http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=21098
Company X
Wholesaler
Supplier
Retailer
# of cases
Salmonella Enteritidis PT14b outbreak 198 of 287 (69%) cases could be plausibly
linked to eggs supplied by one company
Salmonella Enteritidis 14b
National / International
Orange – Austria
Blue
- France
Red
- Germany
Black - U.K
SE PT14b outbreak 198 of 287 (69%) cases could be plausibly linked to
eggs supplied by one company
Key Conclusions
•
•
•
•
MLVA was used to identify this multi-country Salmonella
outbreak
The use of WGS provided confirmation and improved
discrimination between isolates.
WGS data provided a clear link between isolates from
cases, eggs and environmental samples from premises
associated with outbreaks.
This, along with the egg supply network information and
information from investigations in other European countries,
provides compelling evidence to support the hypothesis that
the outbreak was associated with eggs from one Company.
.
Inns et al.EuroSurveill.2015;20(16):pii=21098 http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=21098
E. coli O157 outbreak linked to bagged
watercress
•
•
•
•
•
•
•
•
In September 2013, a national increase in cases of VTEC O157
phage type 2 VT2 was observed
19 cases (14 in England, four in Wales and one in Scotland)
Ten of thirteen cases contacted had consumed watercress
purchased from a major retailer
On 12 September 2013 – first Outbreak
Control Team (OCT) held.
The retailer recalled pre-packed salad products
containing watercress on the same day.
FSA issued a product recall notice to consumers
Trace-back investigations, extensive microbiological testing of
watercress and environmental sampling at farms.
PT8
PT2
NonDomestic
PT2
Domestic
PT21/28
Maximum likelihood phylogeny of
VTEC O157:H7
PT8
Outbreak 1
• We were unable to identify a
PT2
NonDomestic
OB1
PT2
Domestic
OB2
local source for Outbreak 1
• Imported components of the
process were implicated
• These included watercress
seeds from North America and
mainland Europe and peat
from mainland Europe
Outbreak 2
Distinct cluster of cases
and isolate from watercress
irrigation water
PT21/28
Outbreak 2 – Environmental
investigation
Presentation
- edit in Header
and
Footer
Thanks totitle
Caroline
Willis,
PHE
FW&E
Robust SNP Phylogenies for National Surveillance
Porton
Summary of Outbreak
•
•
•
•
The sequencing data identified two separate, concurrent
outbreaks of E. coli O157 PT2 both linked to the consumption of
watercress
Strains from Outbreak 1 cluster were not closely related to any
other UK strain in the Public Health England WGS database
suggesting a possible imported source
Epidemiologically unrelated isolates on the same branch as the
Outbreak 2 cluster included those from human cases in England
with domestically-acquired infection and UK domestic cattle.
Environmental sampling resulted in the isolation of E. coli O157
PT2 from the irrigation water at one of the implicated watercress
farms and WGS showed this isolate belonged to same
phylogenetic cluster as Outbreak 2.
WGS in outbreaks
WGS is good at identifying outbreaks that are:
• Small
• Geographically dispersed
• Spread over a long time and may not have triggered
exceedance
•
•
•
•
•
Time to outbreak detection using WGS is faster than
existing methods
Time to outbreak resolution is faster using WGS.
WGS provides a more accurate outbreak definition
WGS is more robust at calling a case microbiologically
linked or not linked to another case
Provides information on phylogenetic relationships –
ancestral history
Application of WGS to outbreak investigation of gastrointestinal pathogens
Other WGS applications
• Attribution of pathogens to reservoir or
•
•
•
•
source e.g. Campylobacter jejuni and
poultry, cattle etc,
Antimicrobial resistance genes
Virulence gene profile e.g. STEC
Burden of disease – metagenomics to
reduce the diagnostic gap?
Viruses and parasites
Summary
• Area is still developing and likely to see
•
•
•
further changes
Valuable tool for routine surveillance,
outbreak detection and investigation
Still some significant hurdles to overcome
- international collaboration essential
Many other areas where WGS can have
an impact in terms of food safety
Acknowledgements
Public Health England
Kathie Grant, Tom Inns
Food Standards Agency
Jo Edge, Alisdair Wotherspoon
University of Liverpool
Sarah O’Brien
Thank you