An Integrated View on Listeria Genomics and Virulence
Sylvain Brisse
Institut Pasteur
Listeria monocytogenes is everywhere, and
infects humans through food
• ~ 300 human cases/year in France (1,600 in the USA)
• Severity of invasive forms (hospitalization rate around 100%)
• Mortality rate of 20 to 30 % (up to 45% in case of CNS infection)
• Strict control procedures in food industry
Listeria monocytogenes invasive infections
• CNS infection
• Bacteremia
• Materno-neonatal infection
Objectives
Population structure
Strain nomenclatures
Databases
Genome dynamics
Phenotypic diversity
Link genotype-phenotype
Applications: diagnostic, epidemiology, risk assessment…
1
2
3
L. monocytogenes strain typing methods
• Serotyping
• Pulsed-field gel electrophoresis (PFGE)
• Ribotyping
• Amplified Fragment Length Polymorphism (AFLP)
• Multilocus sequence typing (MLST)
• Multi-virulence sequence typing (MvLST)
• Multilocus VNTR Analysis (MLVA)
• Single Nucleotide Polymorphisms (SNP)
• Genome sequencing
L. monocytogenes: four phylogenetic lineages,
of which two are epidemiologically important
4b, 1/2b
1/2a,1/2c
I
II
III
IV
Tsai et al., 2011
Den Bakker et al., 2010
Piffaretti et al., 1989
Konstantinidis & Tiedje 2005
DNA-DNA reassociation
ANI
I
II
III
IV
4.99%
5.3%
7.57%
Average Nucleotide Identity
L. monocytogenes lineages ≈ distinct genomic species
ANI among L. monocytogenes lineages:
Lineage I
4b
1/2b
1/2b
CC1
CC4
CC2
CC3
3b
7 Ragon et al., 2008
Lineage II
1/2a
1/2a
1/2a
EGDe (1/2a)
1/2c
CC7
CC101
CC9
Major L. monocytogenes serotypes ≈ lineages
abcZ bglA cat dapE dat ldh lhkA
3 1 1 1 3 1 3
3 1 1 1 3 1 2 3
1 2 1 7 22 39 7 2
3 1 1 1 3 1 3 1
Alleles Sequence type (ST)
1
1
2
3
3
•Allele • Sequence of one gene (lhkA, 480 bp)
Listeria monocytogenes Multilocus Sequence Typing
www.pasteur.fr/mlst
~3,500 entries, > 1000 STs
> 60 labs
Common language / strains Salcedo et al. 2003
Ragon et al. 2008
MLST: major L. monocytogenes clones
Lineage I
Lineage II
Ragon et al 2008
Chenal-Francisque et al 2011
Haase et al 2014
…
9 14
15
3
9
10
4
1
12
10 10
5
7 5
7
2
7
7
10
1
10
1
13
3
3
12
1
3
14
12
2 3
12
14
3
1
3
8
10
5
9
10
America (n = 77)
Europe (n = 156)
North Africa
(n = 33)
Asia / Middle
East / Oceania
(n = 30)
(No significant Fst)
Chenal-Francisque et al., 2011
MLST clones: Everything is Everywhere CC1
CC2
CC3
Lin. I, others
CC9
Lin. II, others
L. monocytogenes MLST clones are old Haase et al., Env. Microbiol. 2014
CC9
CC11
(ECIII)
CC8 (ECV)
CC7 (ECVII)
CC121 CC101
CC26
CC21
CC155
CC199
CC177
CC16
CC18
CC19 CC193
CC20
CC37
CC2 (ECIa or ECIV)
CC1 (ECI)
CC4
CC3
CC5
(ECVI)
CC59
CC6 (ECII)
CC315
CC195
CC90
ST558
FSLJ1-175
FSLJ1-194 FSLJ2-064
L028
EGD-e
FSLN3-165
10403S
EGD
08-5578, Canada, 2008 TS4/F.6854, USA 1989 J0161, USA 2000 G6054, USA 1994 CLIP 80459, France 1999 H7550, USA 1998-99 J1735, USA, 2002 J0211, USA 2000-2001 Scott A, USA, 1983 Aureli97, Italy 1987 TS38/L.3306, UK & Ireland 1987-89 TS27/L.4738, Canada 1981 TS60/L.4486b, Switzerland, 1983-87 F2365, California, 1986-1987
Outbreak strains:
111
17
1
62
1
2
3
4
> 4
Cantinelli et al., 2013
See also:
Kathariou & coll.
Knabel & coll.
Lineage II
Lineage I
‘Epidemic clones’ ≈ CCs
IIa
IIb
IIc
IVb
Food isolates
2006-2013
Clinical isolates
2006-2013
Food isolates
2006-2013
Clinical isolates
2006-2013
Lineage II
Lineage I
Lineages Serogroups MLST clones
Lineage I
Lineage II
Distribution in food vs. clinical samples
Data from French National Reference Center
NRC
Quantification of MLST clones in sources
2584 clinical
4049 food
6633 isolates
PFGE ApaI, AscI
CC1 CC4 CC6
2005 – 2013
NRC + NRL isolates
M. Maury
Unequal distribution of clones in food and clinical sources
Maury, Tsai et al. Nat Genet 2016
Clinical-associated clones are hypervirulent
Maury*, Tsai* et al. Nat Gen 2016
811 infected patients, immunosuppressive comorbidities
(MonaLisa study, C. Charlier) 6 633 isolates, invasive infections
R2 = 0.9995, P < 0.03 Coll. lab
M. Lecuit
LLS LIPI-1
CC2
CC1
CC6
CC4
CC9
CC31
CC121
Truncation Internal deletion
Absent Present / full length
ST13 CC193
Lineage II
Lineage I
0.01
Variation of conserved virulence factors
Maury*, Tsai* et al. Nat Gen 2016
FA
M000548
F
AM
000450
FA
M003296
FA
M004348
F
AM
003164
FA
M003292
FA
M002725
F
AM
002828
7.0
21.7
29.3
58.2
2.6
0.0
40.7
16.0
20.3
32.2
25.0
35.1
35.9
68.2
46.2
45.0
31.0
52.2
45.2
41.9
37.1
42.5
71.3
58.1
61.5
50.9
65.6
0.01
FA
M002728
FA
M002730
F
AM
002729
FA
M002925
FA
M003293
F
AM
003294
FA
M004349
FA
M003297
F
AM
003298
FA
M003299
FA
M000549
F
AM
001910
F
AM
002113
F
AM
002114
F
AM
002292
F
AM
002293
FA
M003315
FA
M002768
FA
M002772
FA
M004135
FA
M004139
FA
M004150
FA
M004151
FA
M004152
FA
M004153
FA
M004154
FA
M002808
FA
M004136
FA
M004137
FA
M004138
FA
M002769
FA
M002770
FA
M003316
FA
M003317
FA
M003318
20
Present
Absent
Lineage II
Lineage I
Clin
ica
l fr
equ
ency
104 genomes
Novel putative virulence factors
Maury*, Tsai* et al.
Nat Gen 2016
21 Listeria pathogenicity island 4 (LIPI-4)
CC4 (LM09-00558)
1 kb CC4∆PTS
Maury*, Tsai* et al. Nat Gen 2016
Biodiversity of Listeria monocytogenes
• Hypervirulent clones
• Novel putative virulence factors
• First CNS-associated virulence factor
Nat Genet. March 2016
Epidemiology
Diagnostics
Global diversity
Population structure
Ecology (patho)physiology
Integration of public health and biological research
Chenal-Francisque, Maury et al., J Clin Microbiol, 2015
Clonogrouping: rapid multiplex PCR assay
to identify major clones of L. monocytogenes
95.6% of correct identification at clone level
From: [email protected]
Subject: type CT131 Denmark outbreak
Body: Have you observed this type in your country?
Talking the same language:
How can we share nomenclatures of strains?
1. Define genotypic groups and nomenclature designations
2. Develop a nomenclature database
3. Provide on-line genotyping tools
Core-genome genotyping (cgMLST)
Contig 1 Contig 3 Contig 2
Gene 1 …. Gene 2000
~ 2000 core genes
BLAST
~5 days
~30 seconds /strain
~30 seconds /strain
~ few weeks
Standard
codes
A nomenclature proposal for L. monocytogenes strains
Lineage
Sublineage
CT
Lineage - Sublineage – ST – cgMLST type (CT)
LII – SL7 – ST7 – CT932
Nomenclature
database
Provenance data
& genomes
Isolates
databases
Share nomenclature, not genomes or provenance data
Confidentiality of data Standard nomenclature
BIGSdb
Jolley & Maiden 2010
http://bigsdb.web.pasteur.fr/listeria
• L. monocytogenes genotypic categories can be defined at
different phylogenetic depths
• Integration of epidemiological surveillance and fundamental
biology research provides novel insights on pathophysiology,
ecology
• Standard genotyping strategy and public database for strain
nomenclature: to be agreed upon
• L. monocytogenes clones are everywhere; International
transmission both at evolutionary and at epidemiological
timescales
• Need for internationally coordinated surveillance of
L. monocytogenes
Conclusions
Acknowledgements
Applied Maths, Belgium
Hannes POUSEELE, Bruno POT
CDC, Atlanta, USA
Peter GERNER-SMIDT, Cheryl TARR
Heather CARLETON, Lee KATZ, Zuzana
KUCEROVA, Steven STROIKA, John
BESSER
SSI, Denmark
Jonas LARSSON, Eva NIELSEN
Public Health England
Tim DALLMAN, Kathie GRANT
PHAC, Canada Aleisha REIMER, Matthew WALKER, Celine NADON
Oxford University, UK Keith JOLLEY
Funding:
Microbial Evolutionary Genomics, Institut Pasteur
Alexandra MOURA, Mylène MAURY, Marie TOUCHON
Eduardo ROCHA
Center for information Technology, Institut Pasteur
Louis JONES, Emmanuel QUEVILLON
C3BI/HubB
Alexis CRISCUOLO
Julien GUGLIELMINI
Elise LARSONNEUR
Biology of Infection Unit
& National Ref. Center for Listeria,
Institut Pasteur
Yu-Huan TSAI
Vivivane CHENAL-FRANCISQUE
Caroline CHARLIER
Alexandra MOURA
Alexandre LECLERCQ
Marc LECUIT
Top Related