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Genomic Analysis of Meningococcal Serogroup

WXin Wang, Ph.D

Meningitis LaboratoryCDC

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Introduction

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Human commensal bacterium and also one of the common pathogens causing bacterial meningitis.

12 serogroups based on the structure and chemical composition of cell surface associated capsular polysaccharide.

Only six serogroups (A, B, C, W, X, and Y) are associated with most invasive disease.

N. meningitidis

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Frequent horizontal gene transfer events and vaccine-induced immune selection

Antigenically and genetically diverse Dynamic population structure Common typing methods to determine

genetic/antigentic variations Multilocus sequence typing (MLST) Typing of outer membrane protein (OMP) encoding genes (porA,

fetA, fHbp, nadA, and nhbA) Pulse-Field Gel Electrophoresis (PFGE)

N. meningitidis

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MLST Targets the internal region (400-

500 bps) of the 7 house-keeping genes/loci

For each locus, an allele number is assigned to each unique nucleotide sequence.

A sequence type (ST) is defined by the allelic profile of the seven loci.

Strains that have at least four alleles in common are usually defined as the same clonal complex (CC).

Maiden, M., et al. 1998. PNAS

abcZ adk aroE fumC gdh pdhC pgm ST CC4 10 5 4 6 3 8 32 CC32/ET-5 complex

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OMP typingPorA (for porin A)• Class 1 transmembrane protein with 8 loops • Major component of OMV-based vaccines • Two hypervariable regions, VR1 (loop 1) and VR2 (loop 4).• Nomenclature: P1.5-2,16-2• VR1 & VR2 are targeted by the bactericidal antibodies and

are thus under selective pressure.FetA (ferric enterobactin transport)• Formerly FrpB (iron-repressed protein B) • 13 surface-exposed loops• One variable region: F3-3

B vaccine immunogens • FHbp, NadA and NhbA

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Sero-group

Clonal Complex

PorA:FetA

2000-05 2006-10 p value

No. of isolates

Proportion1

No. of isolates

Proportion1

B n=298 n=127 CC41/44 P1.7-2,4:F1-5 17 5.8 2 1.4 0.02 CC32 P1.7,16-2:F3-3 9 3.1 0 0.0 0.02 P1.7,16:F3-3 49 16.6 16 12.7 0.055 C n=277 n=162 CC103 P1.5-1,10-4:F3-9 17 6.1 28 17.3 0.0002 CC11 P1.22-1,14:F3-6 23 8.3 3 1.9 0.009

P1.5,2:F1-30 32 11.6 1 0.6 <0.0001 P1.5,2:F3-6 57 20.6 18 11.1 0.03 P1.5,2:F4-12 3 1.1 28 17.3 <0.0001 P1.5-1,10-4:F3-6 25 9.0 2 1.2 0.002 P1.5-1,10-62:F3-6 0 0 21 13.0 <0.0001 Y n=319 n=196 CC23 P1.5-1,2-2:F5-8 97 30.4 95 48.5 <0.0001 P1.5-2,10-1:F4-1 167 52.4 63 32.1 0.004 NG n=29 n=19 CC32 P1.7,16:F3-3 8.0 27.6 0 0.0 0.03

Strain genotypes with significant changes between 2000-05 and 2006-10,

ABCs

(Wang et al unpublished)

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Significant public health concern due to its high mortality rate world-wide.

Meningococcal meningitis epidemics were reported in many other countries in North America, Asia, Europe and South America.

Shift in the global epidemiology of meningococcal disease in post-vaccine era

Meningococcal disease

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Meningitis epidemics in the meningitis belt

highest incidence (up to 1000 per 100, 000 population) found in the belt

occur in cycles of 8-10 years predominately caused by serogroup A pre-MenAfriVac

http://wwwnc.cdc.gov/travel/yellowbook/2012/

Eradication of serogroup A meningitis epidemics

MenAfriVac, an affordable serogroup A conjugate vaccine developed for African countries

Introduction in Burkina Faso, Mali and Niger in 2010 No serogroup A detected among vaccinated population

http://www.thegatesnotes.com/Topics/Health/The-MenAfriVac-Vaccine-an-Amazing-Success-Story-in-Global-Healthhttp://www.meningitis.org/assets/x/53978

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Post-MenAfriVac Followed the implementation of MenAfriVac,

serogroup W becomes the most prevalent strain causing meningococcal disease in vaccinated countries.

There is an increase in serogroup W disease in the vaccinated countries such as Burkina Faso and Mali.

This is extremely concerning because serogroup W strains currently circulating in Africa may have the potential to cause epidemics and large outbreaks.

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NmW and outbreaks

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The first large NmW outbreak occurred during the Hajj pilgrimage in Saudi Arabia in 2000

More than 300 cases were reported Caused by a strain of CC11 with a PorA type of P1.5,2,

PFGE pattern 40, and 16S type 31 This strain is known as the Hajj clone

Hajj outbreak

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Intercontinental spread of serogroup W (the Hajj clone) since Hajj outbreak: European, Asian, Middle East, North African countries and the United States.

A large outbreak involving nine European countries (the UK, France, the Netherlands, Germany, Finland, Sweden, Belgium, Switzerland, and Norway) was reported immediately following the Hajj outbreak in 2000.

A phylogenetic analysis indicated the Hajj outbreak and associated strains collected from Saudi Arabia, France, Singapore, Finland and the United States in 2000 had identical PFGE pattern (40), 16S type (31) and PorA type (P1.5,2), and belonged to the CC11 (ST-11)/ET-37 complex (ET-27).

Hajj associated outbreak

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NmW strains from 1970-2000 that were not epidemiologically linked to the Hajj outbreak• Diverse genetic background • Some strains were genetically identical to the Hajj clone by PFGE,

MLEE and PorA typing; 16S type differs by 3 nucleotides

The Hajj-related clone circulating in different regions before 2000;

Due to the recombining nature of N. meningitidis, genetic variation occurred during the transmission

The Hajj outbreak led to the global transmission of this clone.

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NmW in the US

No. of Isolat

es

Clonal Complex

Total CC22 CC11 CC23 CC174 CC167 CC168 unassigned95 57 5 7 3 1 1 21

(Wang et al unpublished)

Accounts for <5% of invasive meningococcal disease in the United States

14 W cases in South FL, 2008-9 (6 cases during 2004-7)CC11/ST11, and similar to the Hajj clone by PFGE

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CC11/ET-37 complex has been present in Africa since at least 1993

In 2002, a serogroup W epidemic was reported in Burkina Faso, which affected 12,000 people and caused 1400 deaths.

Caused by a CC11 strain, showing similarity to the Hajj clone

This clonal complex continues to spread in African countries after the 2002 Burkina Faso epidemic and remains to be the prevalent genotype.

Clonal complex 175 emerged between 2002 and 2010.

NmW in Africa

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Clonal Complex No. (%) isolates

CC11 66

CC23 1

CC175 38

NmW strains in the meningitis belt between 2004 and 2010, Pre-MenAfriVac

Caugant DA, Kristiansen PA, Wang X, Mayer LW, et al. (2012) PLoS ONE 7(9): e46019.doi:10.1371/journal.pone.0046019

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Project Objectives

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To determine the genetic similarity and difference of NmW strains from different geographic locations, and the genetic relatedness of these strains to the Hajj clone

To identify genetic markers associated with epidemics and outbreaks.

To determine whether capsule switching has occurred between serogroups W and C.

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Lab_Id Year Country submission type SG ST CCM22702 1998 Algeria endemic W 8814  M23413 2011 Burkina Faso endemic W 11 CC11/ET-37M22803 2005 Chad endemic W 11 CC11/ET-37 M25087 2012 Mali endemic W 11 CC11/ET-37M07293 2000? South Africa endemic W    M23347 2011 United States FL outbreak W    M09261 2002 Burkina Faso epidemic W 11 CC11/ET-37 M09293 2002 Burkina Faso epidemic W    M24695 2010 Burkina Faso epidemic X 181 CC181M22811 2006 Niger epidemic W 2881 CC175

M22831 2009 Niger epidemic A 2859CC5/subgroup

IIIM07149 2000 Saudi Arabia epidemic W 11 CC11/ET-37 M22822 2007 Togo epidemic W 2881 CC175M25419 2005 South Africa source unknown W    M22819 2007 Benin sporadic W 2881 CC175M22740 2001 Cameroon sporadic W 11 CC11/ET-37

M22748 2001Central African

Republic sporadic W 11 CC11/ET-37M22801 2004 Djibouti sporadic W 11 CC11/ET-37 M22471 1993 France sporadic W 22 CC22M22751 2002 France sporadic W 11 CC11/ET-37 M07165 1995 Gambia sporadic W 11 CC11/ET-37 M22189 2007 Mali sporadic W 11 CC11/ET-37M22718 2001 Mauritius sporadic W 11 CC11/ET-37 M22772 2002 Senegal sporadic W 11 CC11/ET-37 M25543 2012 United States sporadic W    

PorA, FetA, 16S, FHbp, NadA, NhbA etc?

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Meningitis Belt Country

Non-Belt African Country

Others

Burkina Faso Algeria FranceBenin Djibouti United State

Cameroon Mauritius Saudi ArabiaChad South Africa

Central African of RepublicGambia

MaliNiger

SenegalTogo

Strain collection

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Strain collection

Submission type Epidemic vs Endemic

Outbreak vs sporadic Serogroup

W A X

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Strain collectionLab ID Year CountryM09261 2002 Burkina FasoM09293 2002 Burkina FasoM24695 2010 Burkina FasoM23413 2011 Burkina FasoM22471 1993 FranceM22751 2002 FranceM22189 2007 MaliM25087 2012 MaliM22811 2006 NigerM22831 2009 NigerM07293 2000? South AfricaM25419 2005 South AfricaM23347 2011 United StatesM25543 2012 United States

26unpublished

Nov 18-28, 2013

Nov 12-14, 2013

March 6, 2013

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Capsular Switching

Allelic exchange of serogroup-specific gene(Swartley et al 1997)

A capsule switching event is defined as an isolate of a certain serogroup belonging to a clonal complex that is commonly associated with a different serogroup.

A possible mechanism to escape the host immune system

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The cap locus of N. meningitidis

B cssA cssB cssCctrBctrD ctrAtex ctrC galEcsb

A csaA csaB csaCctrBctrD ctrAtex ctrC csaD galE

C cssA cssBctrBctrD ctrAtex ctrC cssC csc galE

Y cssB cssC csyctrBctrD ctrAtex ctrC cssA galE

W cssA cssB cssC cswctrBctrD ctrAtex ctrC galE

NG tex galE…other genetic mutations possible

X ctrBctrD ctrAtex ctrC csxA csxB csxC galE

Capsule TransportGenogroup-specific Capsule BiosynthesisGenogroup

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Capsular switching events among the US isolates from 2006-10, ABCs

Serogroup B to C (n=56) Switching from C to B (n=10) Switching between Y and W in both directions

(n=10). Switching from B/C to Y/W (n=4) and Y/W to B/C

(n=7) Switching from B, C, Y and W to other

serogroups (n=9)

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CC11/ET-37 complex is usually associated with serogroup C.

Capsule switching might have occurred between serogroups W and C, but when and in which direction is not known

Capsular Switching between C and W

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Acknowledgement

CDC MVPDB Meningitis Laboratory

CDC MVPDB epidemiology team

National Reference Laboratories (CHUP-CDG, CHU-YO, CHU-SS, and CM, Burkina Faso)

National Reference Laboratory, Mali

Dr. Pierre Nicolas, former WHO CC, France