Comparative genomics and metabolic reconstruction of bacterial pathogens
Genomes of bacterial pathogens and their diversity
description
Transcript of Genomes of bacterial pathogens and their diversity
Genomes of bacterial pathogens and their diversity
Philippe Glaser - [email protected]
1. Introduction: general concepts on pathogenic bacteria and their genomes
2. How to sequence a bacterial genome
3. Two examples: the genus Listeria and Streptococcus agalactiae
Examples of bacterial species and diseases
Tuberculosis Mycobacterium tuberculosis Leper Mycobacterium lepraeCholera Vibrio choleraWhooping cough (coqueluche) Bordetella pertussisSoar throat Streptococcus pyogenes and virusesMeningitis Neisseria meningitidis and other bacteria
Gonococci Neisseria gonorrhoeae Plague (la peste) Yersinia pestisDysentery Shigella flexneriGastric cancer, ulcer, gastritis Helicobacter pyloriMultiple diseases Escherichia coli, Staphylococcus aureus….
Published genome sequence of bacterial pathogens
Shigella 2 Chlamydiae 4+1Escherichia coli 4+2 Neisseria 2Salmonella 3 Branhamella 0 2Helicobacter 3 Bordetella 3Pseudomonas 1+2 Pasteurella 1Yersinia 3 Actinobacillus 0 4Stenotrophomonas 0 1 Haemophilus 2Burkholderia 0 12 Bartonella 3 Flavobacterium 0 1 Legionella 3 Acinetobacter 0 3 Leptospira 2Vibrio 4 Borrelia 1Campylobacter 1 Treponema 2Staphylococcus 4 Mycobacterium 5Enterococcus 1 Rickettsia 3Streptococcus 9 Anaplasma 0 2Listeria 4+1 Coxiella 1Nocardia 0 2 Ehrlichia 0 1Corynebacterium 1+3 Clostridium 2+1Mycoplasma 6 Total: > 80 published genomes
Biodiversity of the microbial worldBiodiversity of the microbial world4 000 000 000 000 000 000 000 000 000 000 bacteria on hearth3,5 billion years of evolution5000 culturable species - 500 000 (?) species
Bacterial diversity in a Yellowstone hot spring
Principle of the experiment:
Sample PCR amplification of16 S RNA
Cloning 300 clones
First analysis by restriction
DNA Sequencingof 122 clones
54 bacterialgroups
84 sequences14 phyla
38 sequences
12 new phyla
(Hugenholtz et al., J. Bacteriol 1998, 180 366-376)
Diversity of the non-culturable bacterial worldDiversity of the non-culturable bacterial world
How to define a bacterial species
• For eukaryotes the species definition is based on sexual reproduction.
Not possible for bacteria1. Phenotypic definition2. Molecular definition:
70% of “similarity” by genomic DNA hybridizationMore than 97% of identities between the 16S RNA genes
=>A convenient definition but not fully satisfactory
Interactions between humans (the host) and bacteria
• The human body constitutes multiple ecosystems for bacterial communities:– The digestive tract– The throat– The skin– Other places are normally sterile (urine, milk, blood)
› Symbiotic bacteria› Commensal bacteria› Pathogenic bacteria• Opportunistic pathogens and obligatory pathogens
Bacteria and their environments
ReservoirAnimalsWaterSoilFood
…
Humanhost
Vectors
The ecology of the pathogenic bacteria or understanding its adaptation to these environments (growth conditions)
Some questions in the study of human bacterial pathogens
• What are the virulence factors and the host - pathogens interaction factors?
• What is the physiology (the metabolism) of the bacteria in interaction with the host?
• What is the evolution of the bacteria which lead to its adaptation to its host, and the relation with the non-pathogenic related species?
• The identification of diagnostic and typing molecular tools
• The identification on a rational basis of antigens for a-cellular vaccines
• The identification of drug targets
How to use genomics (and post-genomics) to solve these questions
Evolution & Biodiversity
Genome variability
DNA repairBarriers to DNA transfer
Point mutationGenome rearrangementGene duplicationHorizontal gene transfer
Selection Biodiversity
=> virulence and pathogenicity=> virulence and pathogenicity
Size of bacterial genomesSize of bacterial genomes
Nanoarchaeum equitans <500 kbMycoplasma genitalium : 0.580 Mb 481 genesMinimal genome 300-400 genesEscherichia coli 4.6-5.6 4289-5648 genesMesorhizobium loti 7.036 Mb 6752 genesStreptomyces coelicolor : 8.667 Mb 7825 genesHuman 3,000.000 Mb 30000 genes
Adaptation : Transcription regulators - vs genome size
(http://www.regx.de/m_project_bioinformatics.php)
Gene transfers in bacteria
Bacteriophages
PlasmidsTransposons
Competence
Transduction
Conjugation
Transformation
Mobile elements and gene gain
• IS elements => no associated function, gene integration by IS mediated homologous recombination, gene inactivation.
• Transposon => carry functional genes• Integron => a platform to incorporate new functions, multi-antibiotics
resistance.• Phages => may carry virulence genes (cholera toxin)• Pathogenicity (functional) islands• Plasmids => may also carry transposons or integrons• + gene duplication
Identification of such elements in genome sequences
Gene lost
• By homologous recombination
• By insertion of IS elements
• By mutation : gene => pseudogene
Evolutionary impact Reductive evolution (M. leprae, Y. pestis, B. pertussis) Role in virulence: lysine decarboxylase in Shigella (cadA+
derivative are less virulent)
Antigenic variation
• By recombination: a gene cassette is inserted in front of an active promoter or remove from this position. (Brucella, Mycoplamsa galisepticum)
• By mutation: variation of a micro satellite sequence length (homo polymer tract) lead to frameshift deletion or reversion (Helicobacter pylori, Neisseria meningitidis)
Protein families and gene duplications
• May arise by gene duplication or horizontal gene acquisition
• Metabolic functions, surface proteins (antigens)
• Correspond to a specificity of a species
• Frequently discovered after whole genome sequencing
Analysis of the genome of a bacterial pathogen
• Annotation of the genome
• Analysis of regulatory genes
• Analysis of inactivated genes (pseudogenes)
• Identification of protein families and mechanisms of phase variation
• Identification of mobile elements
• Identification of atypical regions (recently acquired)
Information obtained from comparative genomics
DNA sequencingDNA sequencing
Two strategies : directed or random
DNA automated sequencing machines produce 800 bases long sequences with an accuracy of 99 %. => How to sequence a 4 Mb bacterial genome with an accuracy higher than 99.99%?
Chromosome Chromosome
Ordering clones of a large-insert library(cosmids, lambda or BAC)
Sequencing clone by cloneof the minimum tiling path
Complete sequence
Random sequencingof a large number of clones
Sequenceassembly
Complete sequence
Directed strategyDirected strategy Random strategyRandom strategy
‘‘Whole genome shotgun’ Whole genome shotgun’
CompleteCompleteGenome sequenceGenome sequence
Chromosome
Large-insert library
(pSYX34 and BAC)
Small-insert library(pcDNA2.1)
closureAnnotation
End-sequencing (small-insert fragments)
End-sequencing (large-insert fragments)
Assembly of sequences in contigs
Organization of a project
Choice of the strategy
Library construction DNA preparation of plasmid clones High throughput sequencing of both ends of inserts
Assembly
Finishing: gap closure and resequencing of low quality regions
Annotation
LibrariesLibraries
Libraries of insufficient quality => No sequence
Important features : coverage of the chromosome, absence of co-ligation, absence of clones without an insert, size of the inserts.
Different types of libraries: * size of the inserts* copy number of the vector
High-copy number vector : 1 to 3 kb inserts 1 to 3 kb inserts Low-copy number vector : 8 to 12 kb inserts 8 to 12 kb inserts Bacterial artificial chromosome : 50 to 100 kb inserts50 to 100 kb inserts
Chromosomal DNA
Nebulization End repair by T4 polymerase
pcDNA: high copynumber vector
Ligation, transformation
Construction of a 1 - 3 kb long inserts library
Recombinant plasmid
TGTGACAC
Two repeated BstXI sites
5’CCAG TGTG ATGG…CCAG CACA CTGG3’3’GGTC ACAC TACC…GGTC GTGT GACC5’
CACAGTGT
Ligation of BstX I adaptors,
Size selection of the inserts
5’pCTTTCCAGCACA3’ 3’GAAAGGTCp 5’
Purification of thedigested vector
(two 5’ protruding ends)
Cloning of DNA fragments of 100- to 300-kb (average, 150 kb) in E. coli
Bacterial artificial chromosome (BAC)
Vector based on naturally occurring F-factor plasmid found in E. coli
» strict copy number control
»stably maintained at 1-2 copies per cell
»lacZ-based color selection of BAC clones with inserts
BAC library construction
Preparation of chromosomal DNA in agarose plugs
Partial digestion with HindIII or BamHI
50 kb100 kb150 kb200 kb
Ligation vector + DNA purified from agarose plugs
Electroporation into E. coli DH10B
Verification of insert size on PFGE gels after NotI digestion
Linearized BAC vector (7kb)
50 kb100 kb150 kb200 kb
Inserts of 70 - 150 kb
AutomationAutomation
High throughput sequencing
DNA Sequencing 15 years ago!DNA Sequencing 15 years ago!
Automated DNA sequencingAutomated DNA sequencing
Automated sequencingAutomated sequencing
Sequence Sequence assemblyassemblyPhred, Phrap, Phred, Phrap, ConsedConsedhttp://www.phrap.org
Statistics and progress of the project
FinishingFinishingRe-sequencing of regions containing low ‘quality’ sequences
Sequencing of ‘missing’ regions
Sequence gapsContig AContig A Contig BContig B
Contig AContig A
Contig BContig B
Contig CContig C
Contig DContig D
Contig EContig E
Contig FContig F
Cloning gaps
Timing of a bacterial genome projectTiming of a bacterial genome project
Library construction and verification (one month)
Plasmid preparation 5000 minipreps per Mb (7 days)
Sequencing : 10000 sequences per Mb (20 days, ABI 3700)
PCR : highly variable (250 reactions per Mb)
Consumable costs : 10 000 Euro per Mb
Listeria monocytogenes
meningitis, encephalitis, septicemia, abortions, neonatal infections, gastroenteritis
foodborne pathogen
dairy products, meat, vegetables, fishTransmission:
Disease:
Population at risk: elderly, newborns, immuno-comprimised,pregnant women
Mortality rate: 30%
Concern for public health Problem for food industry
Ecology of L. monocytogenes
• Ability to survive and to grow in extreme conditions: low temperature, low water activity, broad ranges of pH…
• Ubiquitous in the environment but at very low count
• Variable count depending on the microenvironment and the season at a single location
• Interaction with the vegetal world (silage) and the animal world (waste)
Interaction of Listeria with its hosts
• Carriage is frequent but transient• Low concentration of Listeria in feces• Intracelullar parasite• Ability to cross three barriers: intestinal, hemato-encephalic
and placental barrier • Provokes a broad range of diseases : gastroenteritis,
septicemia, meningitis, encephalitis, abortions• At risk population : immuno-compromised, elderly, pregnant
women and new-born
What are the relations between the two facets of this bacterium?
L. seeligeri
L. innocua
L. monocytogenes
L. welshimeri
L. grayi
B. subtilis
Phylogenetic tree of the genus Listeria
(Pathogenic species)
L. ivanovii
Vaneechoutte et al. Int J Syst Bact. (1998) 48, 127-139
L. monocytogenes
EGDe
L. monocytogenes
4b
L. innocua L. ivanovii
Genome size 2944 kb 2943 kb 3011 kb 2929 kb
rRNA operons 6 6 6 6
CDS 2848 2795 2968 2782
Phages 1 0 5 0
IS 1 (3 copies)
1 transposon
0 0 5
Plasmide -- -- 81.9 kb --
Genome comparison
Listeria monocytogenes
Bac
illu
s su
btil
isL. monocytogenes/B. subtilis synteny
0
500000
1000000
1500000
2000000
2500000
3000000
3500000
4000000
4500000
0 500000 1000000 1500000 2000000 2500000 3000000
L. monocytogenes EGDe
L. i
nnoc
ua
L. i
van
ovii
L. monocytogenes EGDe
Synteny between Listeria genomes
Absence of rearrangement between genomes Rare translocations : probably deletion + insertion
L. monocytogenesL. monocytogeneschromosome mapchromosome map
L. monocytogenes 270 ‘specific’ genes
L. innocua149 ‘specific’ genes
G+Ccontenthttp://genolist.pasteur.fr/listilist
G+C content of the 270 CDSs specific for L. monocytogenes
0
2
4
6
8
10
12
14
25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 52
G+C%
Nb
of C
DS
s (%
)
Total
Specific
Competence operons in L. monocytogenes
comGA B C D E F G
comEA B C
comC
comFA C
27 - 24
37-34 32-21 34-37 30-18 33-18 32-23 31-17
34-3239-6934-44
38-43 35-35
37 : GC%34 : % identities Bs ortholog
2695.1 and 2014.1 two comEC paralogs (DNA binding protein)
0
500
1000
1500
2000
2500
Amino acids
41 surface proteins with an LPXTG motif
InlA-like *= absent from the L. innocua genome
** ** ** ** ** **
****
****
**
**
**
** **** ****
**
**
Sugar PTSHexose phosphate permeaseBile acid hydrolaseArginine deimidaseGlutamate decarboxylase
Metabolic pathways missing in Metabolic pathways missing in L. innocuaL. innocua
Known virulence factors missing in L. innocua
Surface proteins missing in L. innocua
L. monocytogenes / L. inocua comparison
L. monocytogenes - L. ivanovii
L. monocytogenes 345 ‘specific’ genes
L. ivanovii350 ‘specific’ genes
2944 / 0Virulence gene cluster
inlA inlB
inlC
hpt
bsh
The virulence gene cluster
L. monocytogenes
B. subtilisgcaD
prs
ctc
spoVC mfd
yabK
L. innocua
prs ctcprfA plcA hly actA orfAorfBorfZorfXmpl plcB ldh
orfAorfBorfZ ldh ctcprs
orfAorfBorfTorfS ldhprfA plcA hly i-actAmpl plcBprs ctc
L. ivanovii
prs orfB 5,5 kb ldh L. grayi
orfAorfBorfT ldhprs ctcL. welshimeri
prfAprs plcA hly
orfAorfBorfT ldh ctcL. seeligeri
Complex history with several events of insertion and deletions
L. seeligeri
L. innocua
L. monocytogenes
L. welshimeri
L. grayi
B. subtilis
L. ivanovii
L. monocytogenes EGDe
The inlA - inlB region
L. ivanoviilmo0415 amidase inl-like inlA inlB-like inl-like inlB lmo0439
L. innocua
inlA inlB lmo0435 (LPXTG) lmo0439lmo0432lmo0415
17 gènes
wapA-like
lmo0432
lmo0439
Lmo0435 (LPXTG)
lin439
inlA inlBLin439*
lmo0439lmo0432L. monocytogenes 4b
wapA-like
Entrée InlA, InlB
Mouvement intracellulaire ActA
Passage de cellule à celluleActA
Lyse de la double membrane LLO, PlcBLyse de la vacuole
LLO, PlcA
Other virulence genes
L. ivanovii
L. monocytogenes
L. innocua
: pseudogene
hpt, hexose phosphate transport
bsh, bile salt shydrolasebsh2066 groEL
LPXTG groEL
groEL
837 hpt 839295nt 107nt
31nt
L. ivanovii
L. monocytogenes
L. innocua
hpt457nt 153nt
bsh
PrfA box is not conserved
The PrfA box is conserved
Listeria ivanovii - closer to a real pathogen?Some specific functions related to virulence
lmo1240 1241 1242L. monocytogenes and L. innocua
i-inlBi-inlL i-inlKi-inlB2 i-inlGi-inlH
sphingomyelinase-c
lmo1240
i-inlJ i-inlItRNA
L. ivanovii
i-inlF i-inlE
lmo1242
: soluble internalin
Lmo2699 2700L. monocytogenes and L. innocua
L. ivanovii
Lmo2699 2700LPXTG
Capsule biosynthesis ?
tRNA
A second pathogenicity island
And 96 inactivated genes (pseudogenes)
Conclusions Contrary to the rest of the genome, virulence genes have a complex
history. Possible cycle of virulence genes gain and lost. These cycle may play a
role in the evolution of the genus and in the emergence of species. Functions required for intracellular multiplication are conserved between
the two pathogenic species. Interactions with the host and physiopathology are probably different and
involve different factors. The specialization of L. ivanovii is linked to the presence of specific
genes and to the lost of a large number of functions.
What is the diversity within the species L. monocytogenes
Listeria monocytogenes
Epidemiological dataSerovars
• The great majority of human listeriosis cases is caused by 1/2a, 1/2b and 4b strains
• Serovar 4b strains are responsible for almost all major epidemics of human listeriosis as well as for most of the sporadic cases
1/2a1/2b1/2c
3a
3b3c
4a4ab4b4c
4e4d
7
AscI profiles of L. monocytogenes strains -WHO-multi center study
AscI profiles of L. monocytogenes strains -WHO-multi center study
1/2a, 3a
485
388
242
291
145
194
97
48
23
582
kb
Genomic Division I Genomic Division II
1/2b, 3b 4b , 4d, 4e 1/2c,3c
AscI genomic fingerprints of 62 representative Listeria monocytogenes strains
Brosch et al., 1994, AEM 60:2584-92,
• clinical (epidemic) isolates• food isolates• environmental isolates
hybridisation with chromosomal DNA of
Correlation of genomic and epidemiological data
High density membranes for Listeria
New tools for genomic typingNew accurate methods for diagnostics
Development of:
Identification of genes consistently absent or present in e.g. epidemic and clinical isolates
Should allow the:
L. monocytogenes EGDe 1/2a
L. innocua 6a
gene A gene B gene C
L. monocytogenes 4b
control
Hybridization patterns of L. monocytogenes
L.m.sv. 1/2b
L.m.sv. 1/2b
L.m.sv. 4bL.m.
sv. 4b
Hybridized with genomic DNA of:
Hybridized with genomic DNA of:
L.m.sv. 1/2c
L.m.sv. 1/2a
In total 110 strains belonging to all species of the genus Listeria
Listeria ivanovii 5 strains
Listeria innocua 7 strains
Listeria welshimeri 2 strains
L isteria seeligeri 2 strains
L. monocytogenes 94 strains
Serovar: 1/2a, 1/2c, 1/2b, 3a, 3b, 3c, 4a, 4b, 4c, 4d, 4e, 7
Origin: Environment, food, animals, production environnementhuman (sporadic and epidemic cases)
Hybridisation with different Listeria strains
Grouping 460 genes for 112 strains of Listeria
Sérovar: 1/2a, 3a
Sérovar: 4b, 4e, 4d
Sérovar: 1/2b, 3b, 7
Sérovar: 1/2c, 3c
Sérovar: 4a, 4c
Listeria sp.
Lmo0171Lmo0172Lmo0525Lmo0734Lmo0735Lmo0736Lmo0737Lmo0738Lmo0739Lmo1060Lmo1061Lmo1062Lmo1063Lmo1968Lmo1969Lmo1971Lmo1973Lmo1974
ORF2819ORF3840ORF2568ORF1761ORF0029
ORF0799ORF2372ORF2110
L. monocytogenes
I II III
I.1 I.2 II.1II.2
Conclusion
The L. monocytogenes species shows a broad genomic diversity
Genomes are stable and horizontal genetic exchanges are rare.
The species and subspecies are well defined by a set of genes and it seems that there is no continuum between groups.
The notion of species is probably not only an arbitrary one. DNA array is a powerful genome-level typing tool for
epidemiological studies and research.
Streptococcus agalactiaeStreptococcus agalactiae (group B) (group B)
=> Surveillance of pregnant women to avoid mother-infant transmission=> Development of a vaccine ?
Part of the normal flora colonizing the gastrointestinal tract, of an important part of the population, and may colonizethe urogenital tract.
Disease: Rare infections of immuno-compromised adults Leading cause of invasive infections in neonates
septicemia (early onset disease)
pneumonia (early onset disease)
meningites (late onset disease)
Biodiversity within the species S. agalactiae
Characterstics Human Bovine mastitis__________________________________________________
Pigment + -Lactose - +Salicin + +Beta-galactosidase - +Bacitracine sensitivity - +Protein antigens R, Icp X (Finch & Martin, 1984)
Other animal origins: diseases in various mammals and fishes
Human origin: carriage or invasive strains
MLEE, MLST pointed the existence of an hypervirulent lineage.
Two ecovars
Q. What is the genomics basis of this diversity?
S. mutans
S. bovis
S. salivarus
S. pneumoniae
S. mitis
S. sanguis
S. anginosusS. equi
S. pyogenesS. agalactiae
S. uberis
S. suis
S. pleomorphus
Phylogenetic relationship among Streptoccocci
(from Kawamura et al. J. Syst. Bacteriol. 1995)
S. agalactiaeNEM316
S. pneumoniae
2 206 kb
8
2182
2160 kb
4
2236
Size of the genome
Ribosomal operons
CDSs
S. pyogenes
1852 kb
6
1752
Mobile elements
8 IS 12 phage like
integrases2 integrated plasmid
(1 with 3 copies, 42kb)
17 IS4 bacteriophages
105 IS
Genome comparison
SyntenySynteny between S. agalactiae between S. agalactiae and and S. pneumoniaeS. pneumoniae((1141 pairs of orthologous genes)
S. agalactiae
S. p
neum
onia
e
0
500000
1000000
1500000
2000000
0 500000 1000000 1500000 2000000
SyntenySynteny between S. agalactiae between S. agalactiae and and S. pyogenesS. pyogenes ((1170 pairs of orthologous genes)
S. agalactiae
S. p
yoge
nes
0
200000
400000
600000
800000
1000000
1200000
1400000
1600000
1800000
2000000
0 500000 1000000 1500000 2000000
36 recombination breakpoints
14 mobile islands (532 genes)
G+C%
G+C/G-C
Genes related to mobile element within the islands
I
II
III VII VIII
IV
V
VI
IX
X
XI
XII
XIII
XIV
18 kb
16 kb
46 kb
19 kb
11 kb
59 kb
46 kb
46 kb
25 kb
33 kb
86 kb
45 kb
23 kb
int rep tra
int rep rep mob.
Plasm. Phage int
int tnp tnp
tnp
rep parA plasm.tra ssb rep
tra
mob plasm. tra phage rep
tnp tnp pol int rep int rel hel tra rep
Int phage
rep phage int int
int
tRNA-A
tRNA-L
tRNA-TtRNA-R
tRNA-A
tRNA-K
NEM316 - SAG2603 genome comparison
• No chromosomal rearrangement between the two strains
• No integrated plasmid in SAG2603 but three prophages
•1799 orthologs among these two genomes (633 100%
identical)
=> 241 Nem316 genes are missing in SAG2603 (37,
backbone)
=> 258 Sag2603 genes are missing in NEM316 (42,
backbone)
Although highly variable 10 mobile islands are conserved.
NEM316 / SAG2603 - conserved backbone0
gbs0046-47
sga0046
gbs0086-87
sag0086-88
gbs0162-163
gbs0493
cpsJDNMH
gbs1240-1242
gbs1400-1401ABC transporter
sag1330-1331protein R5
gbs1740-1749ABC transporter sag1697-1703
gbs1823His triad prot
sag1780
NEM316
SAG2603
Comparative analysis of island XII
NEM316
98%< <100%95%< <98%90%< <95%80%< <90%70%< <80%60%< <70% <60%
Lactose utilizationLmb scpB
SAG2603 A/B
Mercuric and cadmium resistance
MLST results for S. agalactiae
NEM316
« Hypervirulent »
Sag2603
(Jones et al., 2003 Int. J. Clin. Microbiol.)
adhP : alcohol dehydrogenasepheS : Phenylalanyl tRNA synthetaseatr Amino acid transporterglnA glutamine synthetasesdhA serine dehydrataseglcK glucokinasetkt transketolase
DNA arrays hybridization for genome characterization
• 10 invasive ST-17 strains (MLST study)• BM110, hypervirulent clone defined by
MLEE• 18 invasive strains (Hôpital Necker)• 13 carriage strains (Hôpital Necker)• 14 strains from bovine mastitis• 12 strains of animal origin (horse, dog, cat,
rabbit, guinea pig, fish)
68 strains analyzed by MLST and hybridization
Genome diversity is essentially located within genomic islands
0
50
100
150
200
250
300
inv_b9inv_h11inv_j81inv_j95inv_mk2inv_wc3inv_1000inv_1572inv_1002inv_1560inv_1568inv_318inv_1573port_38bisport_41bisport_37.39port_65.8bisport_60_36bisbov_411.07bov_527.25bov_543.05bov_547.25bov_549.13bov_501_19bov_44poisson_2_22chat_3448_97chat_693chien_928662gui_pig_622lapin_6144_98
islandsbackbone
st17st1st10,6,9
st23st23 st103
Hierarchical clustering of 69 strains and comparison with MLST data
st19
Two loci heterogeneously distributed among isolates
rofA hemagglutinin Glycosyl transferase secY secA
rofA and rogB are mutated in sag2603
fibronectin binding protein srtB srtCLPXTG LPXTGrogB
I
II
I ------------------++++++++++++++++++++++++++++++++--++++++++++II ------------------++++++++++++++++++++++++++++--------+-+-++++
Conclusion
• Strains from different origin do not cluster except invasive ST17 strains.
• ST17 strains constitute a highly homogenous group
• Diversity reside mostly within islands
• Antigenic diversity is highlighted by genome analysis and is
found both within and outside islands
• DNA arrays, a powerful method for molecular epidemiology