Diversity of Genome Structure in Salmonella enterica Serovar Typhi Populations
Whole genome microbiology for Salmonella public health microbiology
-
Upload
philip-ashton -
Category
Science
-
view
1.876 -
download
4
Transcript of Whole genome microbiology for Salmonella public health microbiology
Revolutionising Public Health Reference Microbiology Using Whole Genome Sequencing: A Case Study with SalmonellaPhilip AshtonBioinformatician, Gastrointestinal Bacteria Reference Unit
2 Salmonella WGS for Public Health
PHE are using WGS as the routine test for Salmonella
reference microbiology.
All Salmonella isolates are being sequenced.
No routine serotyping.
Why?How?What?
Salmonella WGS for Public Health3
4 Salmonella WGS for Public Health
Why do we want to implement WGS for Salmonella
reference micro?
5 Salmonella WGS for Public Health
> 38000 cases of Salmonella
gastroenteritis in the UK per year, of
which > 11000 report to their GP.
Tam et al, 2011, Gut gut.2011.238386
Typhoidal Salmonella as well.
77 deaths in UK 2008, FSA Foodborne
Disease Strategy
6 Salmonella WGS for Public Health
Tom Innes & PT14b OCT
‘protecting and improving the nation’s
health’.
7 Salmonella WGS for Public Health
Previous typing method - serotyping
Satheesh Nair
8 Salmonella WGS for Public Health
David Powell
9 Salmonella WGS for Public Health
PHE Salmonella surveillance data - http://bit.ly/1CdEOe0
10 Salmonella WGS for Public Health
PHE Salmonella surveillance data - http://bit.ly/1CdEOe0
Salmonella is a major public health issue.
Existing typing/subtyping has limited phylogenetic insight.
Need higher resolution typing methods (WGS) for monitoring trends and detecting outbreaks.
Unique opportunity for global data sharing
11
Summary – why WGS?
Salmonella WGS for Public Health
12 Salmonella WGS for Public Health
How will WGS based Salmonella reference microbiology work?
WGS implementation at PHE2011- 2015
2 MiSeq machines 2 HiSeq 2500 high-throughput machines
Capacity > 3,000 genomes per week+
PHE investment in WGS: financial, laboratory, bioinformatics, data
handling, staff training
=
Salmonella WGS for Public Health13
Identification / Mixed – kmer gateway
K-mer Gateway
14
AcetobacterAcinetobacterActinomycesAeromonasAggregatibacterBacillusBacteroidesBartonellaBifidobacteriumBordetellaBorreliaBrucellaBurkholderiaCampylobacterChlamydiaChlamydophilaClostridiumCorynebacteriumDesulfovibrioEnterobacterEnterococcusEscherichiaFrancisellaFusobacteriumGardnerellaGordoniaHaemophilusHelicobacter
KlebsiellaLactobacillusLegionellaLeptospiraLeuconostocListeriaMorganellaMycobacteriumMycoplasmaNeisseriaNocardiaPaenibacillusPrevotellaPropionibacteriumPseudomonasRhizobiumRhodococcusRickettsiaSalmonellaShewanellaShigellaStaphylococcusStreptococcusStreptomycesTreponemaUreaplasmaVibrioYersinia
Off all the k-mers of length 18 in each reference genome what percentage are in our sequencing reads?
Can be used to identify cross species contamination.
Salmonella WGS for Public Health
99.7% accurate subspeciation
15 Salmonella WGS for Public Health
Serotype inferred via MLST
Satheesh NairAchtman et al., 2012
MLST to infer serotype
16
Method
Short Read Sequence Typing
ST (and eburst group)
Serotype
Result
Inouye et al., 2012
6887 strains with WGS and pheno
6616 (96%) results matched
Salmonella WGS for Public Health
2 serotypes – 1 ST/EBGLab errorNo ST/serotype lookup
Novel Sequence Types
17 Salmonella WGS for Public Health
SISTR/SeqSero
18 Salmonella WGS for Public Health
Challenges:• Many different eburst groups (of STs) – have to be analysed separately• Hundreds of strains a week• Rapid, hands-off analysis
Solution – SnapperDB:
Most common serotypes – SNP typing
SampleFASTQs(with ST)
EBG 1 - Typhimuriumdb
db
db
db
db
EBG 4 - Enteritidis
EBG 13 - Typhi
EBG 3 - Newport
EBG 11 – Paratyphi A
…
30 mins - parallel 5 min – 1 hour
Tomorrow – pipeline se
ssion
19 Salmonella WGS for Public Health
What will be the output of WGS for Salmonella
reference micro?
• Example of use in outbreak – PT14b• Strain characterisation – antibiotic
resistance profile• Future perspectives
20 Salmonella WGS for Public Health
Salmonella NGS at PHE
Salmonella Enteritidis Phage Type 14b
International outbreak
Tom Inns/PT14b Outbreak Control Team – Inns et al., Eurosurveillance, 2015
22 Salmonella WGS for Public Health
Tim Dallman, Tom Inns, Thibaut Jombart
Hierarchical clustering and Geography
23
Alison Waldram & Gayle Dolan
24
Phenotype S Phenotype R Error%Genotype S Genotype R Genotype R Genotype S
CHL 580 2 39 3 0.78SUL 466 1 149 8 1.40TET 467 3 151 3 0.93TMP 562 3 57 2 0.78AMP 484 5 134 1 0.93CTX/CAZ 618 1 5 0 0.16CPR 619 1 4 0 0.16FOX 612 0 7 5 0.78CIP 478 4 138 4 1.25NAL 485 2 127 10 1.87GEN 608 1 14 1 0.31TOB 613 2 8 1 0.47AMK 622 0 1 1 0.16STR 486 18 119 1 2.96 7700 43 953 40 0.95
AMS: Genotype vs phenotype
Major errors Very major errors
Michel Doumith/Martin Day
Validation : 642 Salmonella strainsResistance : 57.5 % susceptible 24.7 % multi-resistant (> 2 classes)
Salmonella WGS for Public Health
Future perspectives
25 Salmonella WGS for Public Health
• Move from MLST to kmer/phylogenetic placement for sorting before SNP analysis, especially for Enteritidis.
• MinION sequencing: • Outbreak investigation (Quick, Ashton et al., Genome
Biology, 2015, v16, p114), • Solving complex + interesting regions of the genome
(Ashton, Nair et al., Nature Biotech, 2015, v33, p296)• Live streaming of sequencing data – google ‘Loman
labs’• National (One Health) and international (GenomeTrakr)
real-time collaborations
Summary• WGS is a single test – multiple assays• Better quality information than existing
tests• Evolutionary and phylogentic insight
Salmonella WGS for Public Health26
Acknowledgements
27 Salmonella WGS for Public Health
MicrobiologyElizabeth de Pinna, Tansy Peters, Satheesh Nair, Martin Day, Anna Lewis, Tim Dallman, Kathie Grant and other staff in the Salmonella lab
EpidemiologyAlison Waldram (FETP), Richard Elson, Chris Lane, Tom Inns
Genomic Services Unit Cath Arnold and team
Bioinformatics UnitJonathon Green, Anthony Underwood, Rediat Tewolde, et al.